Small Things Considered

And The Winner Is…

by Elio

The transposases! The contest, in case you wonder, was for the most abundant set of genes in the known universe, or at least in the genomic data banks available on Earth. Aziz, Breitbart, and Edwards examined some 10 million genes from 2137 sequenced genomes (47 archaeal, 725 bacterial, 29 eukaryotic and 1336 viral) plus 187 metagenomes, in search of the gene set that is both most abundant and most ubiquitous. (Ubiquitous refers to genes that carry essential functions and are thus needed by every genome. A gene that is highly abundant in some ecosystems but not ubiquitous is likely to play an important habitat-specific role.) The winners, by far, were the genes for transposases. One could squawk that the genomes selected for sequencing to date reflect a bias towards “germs” and other microbes of human interest, and that so far the source environments for metagenomes represent only a small sample of what’s out there. But given the off-scale preponderance of the transposase genes, there is little to make one question their dominance.

Abundance of different functional roles in 2137 genomes plotted against the ubiquity of these
functional roles (defined as the number of genomes in which the functional role is represented
at least once). Source.

Not everybody seems to have transposases; only 1/3 of the viruses in this study have them. But those organisms that do tend to possess multiple copies—close to 40 on average. If you take a set of 2000 randomly sampled genes (the number found in a typical bacterial genome), 22 will encode transposases. They are also more common in free-living bacteria than in obligate pathogens or endosymbionts. The champion transposase-carrier? Possibly the marine cyanobacterium Crocosphaera watsonii, which has more than 400 copies.

So, why transposases? The authors point to their general usefulness to both viruses and cellular organisms in promoting the spread of mobile elements, thus leading to diversification. “Selfish genes” indeed! As the authors write, By securing their own replication and dissemination, transposases guarantee to thrive so long as nucleic acid-based life forms exist. This rather implies that they might imagine non-nucleic acid based life forms exist somewhere. Who says that bioinformaticists are just computational nerds?

Aziz RK, Breitbart M, & Edwards RA (2010). Transposases are the most abundant, most ubiquitous genes in nature. Nucleic acids research PMID: 20215432

When Crenarchaeota Divide, They Multiply

by Jenn Tsau and Suzy Szumowski*

Sulfolobus acidocaldarius, an extreme thermophile occurs in
geothermally-heated acid springs, mud pots and surface soils;
it can withstand temperatures from 60 to 95 ºC, and a pH of
1 to 5. Left: Electron micrograph of a thin section (X85,000);
Right: Fluorescent photomicrograph of cells attached to a
sulfur crystal. Source.

When cells divide, it’s really important that they do so equitably so you don’t end up with some having too much and some too little cell material; the results of either could be deleterious. Therefore, over the billennia that life has been around, mechanisms to carefully divvy up intracellular goodies like DNA and cytoplasm have evolved. You can imagine that interest peaked in those who pay attention to such things when it was found that a group of microbes belonging to the Crenarchaeota lack any of the known cell division machinery. Crenarchaeota are a kind of Archaea that includes species known to survive in extremely hot or cold environments. And when we say extreme, we mean deep arctic-cold and volcano-hot—literally. Other known forms of life have some variant of actin (known as MreB in bacteria) or tubulin (aka FtsZ), which are involved in cell division or maintenance of cell shape. Until late 2008, when a pair of papers was published, no such proteins were found in the crenarchaea and their division remained quite mysterious.

Regardless, these hardy little archaea are getting the job done, one way or the other. It has been known for some time that the Crenarchaeota possess homologs of a system called ESCRT (endosomal sorting complex required for transport). This is an attractive candidate for managing division in these organisms. Why? Because the ESCRT system helps detach daughter from mother cell during cell division in metazoans—such as insects, octopuses, and elephants. So it seemed plausible that these proteins might be helping here as well.

Sulfolobus acidocaldarius. Green: labeled antibody
specific to the Vps4 (top) and ESCRT-III (bottom)
proteins. Red: membrane staining. Blue: DNA staining.
Merged image. Source.

With this starting point, the authors of a recent paper began investigating a possible role for ESCRT proteins in the cell division of a fairly well-studied crenarchaeon, Sulfolobus acidocaldarius. Using PCR and Western blotting, they monitored the levels of ESCRT-like proteins found in synchronously dividing S. acidocaldarius and found that the levels of two such proteins (ESCRT-III and Vps4) were enhanced in dividing cells. Using immunofluorescent staining, both of these two ESCRT components were found at the midline of dividing cells. The regulation of both appeared to follow the same pattern, thus it seemed possible that they interact physically. To figure out if this was the case, the authors used a yeast two-hybrid system to isolate the specific domains of each protein that were binding the proteins to each other. With this information, they postulated what this interaction might look like at the structural level. As you can see, ESCRT-III lies flat snugly against Vps4, which is its binding partner.

A schematic representation of the interaction of
ESCRT-III (Saci1373, in red) with the Vps4 (Saci1372
MIT domain in yellow). Source.

To seal the deal, they needed to show that removing or altering these components would affect cell division did just that through additional immunofluorescent staining experiments. When they grew cell lines expressing a nonfunctional form of Vps4 with a defective ATPase domain, they found some normally-sized cells but many gargantuan cells, the latter being unable to divide and containing abnormally high amounts of DNA. This implies that ESCRT and Vps4 are key players in Sulfolobus cell division.

This novel function ascribed to ESCRT proteins paints a more complete picture of cell division throughout all the domains of life. We can all rest a little bit easier knowing that there is a logical explanation for the continued existence of Crenarchaeota on this earth, even though they do not possess the better known prokaryotic cell-division proteins.

Jenn (left) and Suzy (right) are students in the University of California at San Diego/San Diego State University Integrative Microbiology graduate course during the 2010 winter quarter.

Fine Reading: March of the Microbes by John Ingraham

by Elio

A good friend has written a good book. March of the Microbes, just published by Harvard Univ. Press, presents tales from the microbial world, wide and deep. It emphasizes microbial activities that you can see, smell, touch, taste, and, if you include cows belching, hear. John calls these encounters sightings. Some are manifested in our daily doings, such as the teeth we brush, the bread we eat, the beer we drink, or the fish we left overlong in the ‘fridge. Others require us to go farther afield, making trips to the hot springs of Yellowstone Park, fields of legumes, munition factories, or the San Francisco Bay. You may have heard (or seen) many of the sightings described, although some are not commonplace. Did you know that starch becomes the ubiquitous high fructose corn syrup via degradation by bacterial enzymes? Or that the famed Carlsbad Caverns are the result of bacterial mining? The microbiologist will find the stories both illuminating and authoritative. The lay person will be astonished at the marvels of our unseen world.

This is a book written with love. John is passionate about all microbes he has ever encountered, bar none, and shares his fervor in a clear, unpretentious manner. I should know. Along with Fred Neidhardt, we have our names on four (or is it five?) books.

All Is Fair in Love and Warfarin

by Shigeki Miyake-Stoner and Spencer Diamond

Warfarin, up close and personal. Source.

It turns out the drug warfarin has nothing to do with war, but it does involve a recently discovered link between bacteria and humans. Warfarin derives its name from WARF, the Wisconsin Alumni Research Foundation where it was first developed, and -arin from its coumarin-like structure.

Warfarin is used in two roles: to kill pests, and to save human lives. It was first designed for use as a rodent poison, an anticoagulant that would cause mice and rats to gradually hemorrhage and bleed to death internally. Some years later, it was realized that this drug would be useful for humans too! Patients susceptible to dangerous blood-clotting or thrombosis can take appropriate amounts of this drug to "thin" their blood. Warfarin works by inhibiting Vitamin K epOxide Reductase (VKOR), an enzyme that recycles vitamin K after it acts as a cofactor in activating clotting factors. Now you might ask, what does this have to do with bacteria? Well it turns out bacteria have more in common with mammals than we thought!

Source.

A VKOR homolog was identified in Mycobacterium tuberculosis (Mtb), the bacterium responsible for human tuberculosis. Why might this bacterium have a VKOR? Surely it doesn't need to regulate its blood clotting. Mycobacterial VKOR (MtbVKOR) is involved with making disulfide bonds in proteins, and is quite important for the bug's life processes. A recent paper by Dutton and colleagues investigated the role of MtbVKOR in bacteria.

Knocking out the gene encoding MtbVKOR severely affected growth of Mtb. So did growing Mtb in the presence of warfarin. The authors had previously noted that adding this gene to E. coli restored flagellar function in a mutant lacking the cell-envelope protein DsbB. Since they knew that DsbB is an enzyme involved in forming disulfide bonds, they performed experiments to see if MtbVKOR had the same activity. Indeed, both enzymes mediated the same electron transfer step involved in disulfide bond formation in the E. coli flagellar protein FlgI. Notably, these same transgenic E. coli grown in the presence of warfarin lost both MtbVKOR's disulfide activity and their motility.

SEM of Mycobacterium tuberculosis. Source.

To further compare VKOR and MtbVKOR, warfarin resistant mutants of MtbVKOR were sequenced and found to have point-mutations similar to those that appear in humans resistant to the drug and who require higher dosage.

Although found in vastly different organisms, VKOR and MtbVKOR have the same basic function. The downstream effects of their activity are definitely divergent, but both can be inhibited by the same small molecule, warfarin. It is now clear that this enzyme can be studied for the development of better anticoagulants and better tuberculosis treatments.

The story of warfarin should be of interest to drug companies. In an age of blockbuster drugs and a pill for just about anything, one would think drug companies are doing rather well. Over 12 million Americans take cholesterol-lowering statins, and practically everybody you know has taken at least one prescribed pill in the past year. However, drug companies are begining to feel a pinch as many of them have put all their eggs in one basket, relying on one best-selling drug or another. Many of them are now seeking to expand the list of conditions that can be treated with each one of their drugs. By doing so, the companies can sell more of their drug while it is still patent protected. The evolutionary conservation of enzymatic function and structure will likely lead to more warfarin-like examples, cases where we can find new tricks for an old dog.

The bottom line is that all of this crosstalk is rooted in evolution. When change proceeds without design, as in evolution, it is not easy to create something complex from scratch. We often see that the useful products of evolution get re-used and modified throughout time.

Shigeki (left) and Spencer (right) are students in the University of California at San Diego/San Diego State University Integrative Microbiology graduate course during the 2010 winter quarter.

Talmudic Question #59

Which prokaryotic cells are more abundant on Earth: planktonic (free living) or sessile (adhering to surfaces)? Keep in mind that the ocean waters contain abundant floating surfaces, e.g., marine snow.

On the Continuity of Biological Membranes

by Franklin M. Harold

The complex world of cell membranes. Source.

Thirty years ago, Günter Blobel of the Rockefeller University published a short paper entitled Intracellular Protein Topogenesis, which laid the conceptual foundations for our understanding of how cells build membranes. To serve their functions, peripheral and integral proteins must be inserted into the right membrane with the correct orientation, and most of the article focused on the manner in which this may be achieved. But it also underscored two startling implications of the proposed procedure: first, that every membrane must be derived from a pre-existing membrane; and second, that all extant biological membranes are descendants of the plasma membrane of the first primordial cell.

Blobel’s article became a classic, and spawned a small industry concerned with the molecular mechanisms that target proteins to the recipient membrane and then either translocate or insert them. In a nutshell, the information that specifies a nascent protein’s disposition is contained in its sequence. One segment of that sequence recognizes a receptor protein embedded in the target membrane, commonly part of the translocon; other segments specify whether the amino acid chain is to be taken clear across the membrane or inserted, and with what orientation. Membrane proteins may be processed concurrently with their translation, or after their production is complete. In prokaryotic cells the proteins are produced and handled directly; in eukaryotic cells they are first inserted into the membrane of the endoplasmic reticulum, and then transferred to their target membrane by cargo vesicles. The details can be found in textbooks of molecular cell biology. What concerns us here is the inference that membrane heredity is a fundamental principle of biology. A functional membrane, studded with a particular set of enzymes, transport carriers and receptors, can never be generated de novo; it must arise from a pre-existing membrane, either by modification (for example, the membranes that surround bacterial spores) or else by growth and division or vesiculation. Moreover, since proteins will only be inserted after interaction with a complementary receptor (and that includes the receptor protein itself), a growing “genetic” membrane propagates its own kind.

The idea that membranes are inherited was by no means novel in 1980; cytologists had been musing on it for two decades. But it was quite another matter to assert that it must be so, that “omnis membrana e membrana.”

The Next Generation (Or Two)

by Elio

Source.

Student blogs there are that gladden an old man’s heart. Here’s a sampling.

In Catalogue of Organisms, Christopher Taylor, a student of arachnids in Perth, Australia, posted a new interpretation of the mysterious Prototaxites—giant, 8 meter tall fossils some 400 million years old that predate any plants of that size. It was thought that these megastructures were fungi (see our post on this). It has now been proposed that they are sheets of liverworts that rolled up as they cascaded down slopes. Christopher points out things that may be wrong with this scheme.

In Skeptic Wonder, Psi Wavefunction, an undergraduate in British Columbia, takes on the term “Oncogene” and explains why it should disappear forever. Her writing is so lively that we published a guest post by her recently.

In Micro Writers (“written by students to students”) that comes to us from Cairo University, Mariam points to the wisdom of escaping from anthropocentric to biocentric microbiology. This post is based on a commentary by Ramy Aziz published in Gut Pathogens that was highlighted on our blog not long ago.

In Extreme Biology, students post about "anything biology-related." These students have not yet graduated from high school! To our delight and awe, Amy Ciardiello, a 9th grade violinist, writes about "violin-making and fungi"—a topic we had previously posted (here and here) on STC. She accompanies her post with a superb performance of the 1st movement of Haydn's Concerto No. 2 in G Major. Go there and feast both your mind and your ears.

We welcome notices of other microbiological research blogs presented by students.

Mother’s Love

by Elio

Binary fission is a most impressive invention. In one fell swoop, it ensures that progeny cells are born alike and endowed with the same potential for growth and survival. Simple as it sounds, it must have taken considerable evolutionary contortions to make it function so well throughout the living world. But there are cells that have adopted an alternative mechanism, where cell division is asymmetrical, where one progeny cell is made from a “mother cell” that keeps generating “babies.” The best known example is, of course, budding in yeast. But other cells also arise in this fashion, including some bacteria, the sexual spores of mushrooms, and even some plant cells.

The polarisome in a forming yeast bud. Actin cables
growing out from the polarisome transport protein
aggregates to the mother cell. Source.

So, is there an advantage to bypassing binary fission and budding instead? It would seem that way. Recent work from Tom Nyström’s lab has shown that proteins that become damaged in the course of cell growth flow back into the mother cell and leave the young bud free of such impediments. The damage to proteins is often due to oxidation by reactive oxygen species. Damaged proteins tend to form aggregates. This can be bad, getting rid of them is good. How do these proteins accumulate in the mother cell? Interestingly, protein aggregates hitch a ride on actin filaments that grow from the growing bud to the mother cell. Such filaments are assembled at the tip of the bud in a structure the authors call a “polarisome,” which is made up of core proteins plus some involved in actin polymerization (formins). Also required is the age-retardant deacetylase, a protein called Sir2 and aka sirtuin. Previously known as a life span modulator—not just in yeast but in worms, fish, and mammals as well—Sir2 has now been found to also be involved in actin-related processes, hence in polarisome formation. It gets more complicated. For an overview, we suggest a commentary by Guarente.

(A) Exponentially growing young yeast cells to compare their size and
morphology to that of old cells. (B) A typical lifespan determination.
The number of budding cycles that each of a set of 50 ‘virgin’ cells
undergoes before it stops dividing was determined by micromanipu-
lation and counting of budding cycles. (C) M is the terminal mother cell
after 15 cell cycles. Note the enormous size as compared to young cells
and the surface changes. D14 is the second but last daughter that did
not completely separate from the mother and did not give rise to new
living cells. Source.

Now let’s look at this story in a broader context. It’s not just about shipping dirty laundry off to mother. One consequence of the asymmetry of budding is that the mother cell retains its bodily integrity bud after bud, whereas this is lost when a cell divides by binary fission. In yeast, a mother cell can bud some 15-30 times, after which it conks out. How do we know? Counting the number of times a cell gives off buds is done patiently under the microscope, using a micromanipulator to remove each daughter cell once it separates from the mother cell. Keep this up until the mother cell gives off no more buds. Imagine teasing the new cells away for the mother cell 30 times or so! (This seems to have been done first in 1950 by A. A. Barton, working for a British brewing company.) This phenomenon is called senescence, and is visually illustrated by wrinkles of old age and the unusually large size of the Grande Dame. Newly formed cells start the process anew, each daughter cell becoming a mother cell of its own. However, in time, the newly made cells are less competent for further budding. Not surprisingly, yeast is a favorite for studies of cell polarization and its possible role in senescence. Many papers have been written on this subject.

A favorite connection between yeast budding and aging relies on a 120-year-old theory by August Weismann. He postulated that aging evolved from the need to separate germ cells from somatic cells. Germ cells need to be protected from damage; somatic cells can ”take it.” One of the reasons given is that additional resources must be bestowed on germ cells to ensure their genetic stability. Somatic cells, in contrast, do not have such mechanisms and thus accumulate damage.

This is one way to think about asymmetric cell division. Formally speaking, the mother cell acts as a somatic cell that produces multiple germ cells, the buds. Each, when grown, becomes a mother cell with full reproductive potential, able to produce a full complement of buds of her own. During budding, the young bud escapes from the cell damage represented by the aggregated proteins, thus foiling at least that aspect of cell aging.

If asymmetric cell division affords such protection of the germ line, why don't all cells do this? The question is not terribly relevant to multicellular somatic cells, given that they are not involved in germ line propagation (unless some investigator teases their nuclei out and introduces them into eggs). But how come more unicellular microbes haven't adopted the budding strategy? That is a question for another time.

Given that yeast is the best known of all eukaryotic organisms, allowing endless kinds of genetic manipulations, it’s no wonder that it has become a model for the study of aging. And here I thought that I would be a good subject for researching what happens in old age!

Addendum: As Qetzal noted (see his comment below), I could have mentioned that a possibly analogous phenomenon has been reported for bacteria. When E. coli divides, the "old" cell pole accumulates chaperones involved in the aggregation of (presumably) damaged proteins. Eventually the "old" cells lose their reproductive capability. Something analogous takes place in Caulobacter crescentus. Thus, bacteria may also use the strategy of segregating damaged proteins in aging cells, to the advantage of the population as a whole.

Liu, B., Larsson, L., Caballero, A., Hao, X., Öling, D., Grantham, J., & Nyström, T. (2010). The Polarisome Is Required for Segregation and Retrograde Transport of Protein Aggregates Cell, 140 (2), 257-267 DOI: 10.1016/j.cell.2009.12.031

Prophage Masquerade

by Merry

Source.

Roseovarius nubinhibens recently joined the exclusive club of about a thousand bacteria whose genomes have been sequenced. Why this honor? It’s a member of one of the most ubiquitous and most intensely studied clades of α-Proteobacteria, the marine roseobacters. This populous group participates in important jobs, including the global cycling of sulfur, climate regulation, and even modulation of our day-to-day weather. (For the latter, see our earlier post.) To appreciate the sulfuric importance of R. nubinhibens in particular, we need to begin with the major quantities of dimethylsulfoniopropionate (DMSP) made by marine algae for whom it serves as an osmolyte. Roseobacters then enter the picture, their work being to break down the DMSP by either of two pathways. One route converts DMSP into volatile DMS that can give rise to sulfate aerosols that act as cloud seeds. The other pathway leads to methanethiol, a primary sulfur source for marine bacteria. The relative amount of sulfur that flows each direction matters. R. nubinhibens was one of the first bacteria found to be able to carry out both conversions, thus is a potentially important switch point. The researchers reporting its isolation (José González and colleagues in Mary Ann Moran's lab) dubbed their new species Roseovarius nubinhibens (from the Latin nubes, for clouds, and inhibens, for inhibiting).

Virus-like particles generated by mitomycin C induction of R. nubinhibens
ISM. Siphovirus particles are commonly seen in the induced lysate, and
many of them appear to be “empty.” Bars = 50 nm. Source.

The R. nubinhibens genome has been sequenced. Any prophages on board? A routine search using Prophage Finder (software that looks for clusters of phage-related genes) turned up several candidates. Let's refine the question and ask whether there are any inducible prophages on board. Add mitomycin C (a potent DNA cross-linker) to a growing culture and wait 24 hours. Voilà! You get ~10¹⁰ VLPs per ml compared to ~10⁵ for control cultures. (What is a VLP? It's a Virus-Like Particle, i.e., something that looks like a virus but hasn't been shown to infect like a virus.) The induced VLPs have the long, flexible tails and polyhedral heads characteristic of siphoviruses, but most of these VLPs are empty-headed and/or have broken tails, suggesting that many are defective.

Was one of the predicted prophages the source of the VLPs? The DNA strands inside the VLPs were shown by pulsed-field gel electrophoresis (PFGE) to be about 30 kb, suggesting that the prophage might be about the same. None of the predicted prophages were near this size. So the researchers then searched the genome for any phage-related genes. They were lucky. They found an integrase gene along with a few other recognizable phage genes in a 27 kb region that Prophage Finder had overlooked. They concluded that they had found the prophage associated with the VLPs because they identified a gene in this stretch of DNA that encodes a major capsid protein and showed it to be identical to the capsid gene in the VLP DNA.

So what else does that prophage DNA encode? A few genes required for the prophage life cycle could be tentatively identified. Of the 28 genes that had significant hits in the GenBank database, 25 are most similar to genes of unknown function previously found in Rhodobacterales bacteria. Might those 25 "bacterial" genes actually be phage genes that are part of the unsequenced majority and that are residing in prophages? No wonder Prophage Finder—looking as it does for known phage-related genes—didn’t find the real prophage! Since more than 99.9% of phage diversity has not been captured in any database, hunting for prophages with Prophage Finder is like going birding with a field guide that includes less than one species out of a thousand.

Other sequenced members of the Rhodobacterales harbor related prophages, suggesting that this type of prophage is common and may be active in horizontal gene transfer (HGT) among these bacteria. Tantalizingly, there may be a subplot to this story, with even more HGT going on. The PFGE analysis of the VLP DNA also found a few short segments of 3, 4, and 12 kb. The authors suspect that these may be random bits of host DNA packaged inside capsids, i.e. gene transfer agents (GTAs). (For our earlier post about GTAs, click here.) GTAs transfer genes between members of the same species. Their production is controlled by the bacteria, and neither viral infection or cell lysis is involved. GTAs have been found only in α-Proteobacteria, with a particularly high incidence among the roseobacters. Previously, 21 of the 22 sequenced members of the marine Roseobacter clade had been found to carry the genes for GTA production; now R. nubinhibens makes it 22 out of 23.

What do GTAs have to do with prophages? Did one evolve from the other? If so, in which direction? In the words of two researchers in this field: The overall pattern of relationships between these various GTA, phage and prophage elements supports the notion of a continuum of sequence relationships resulting from transfers throughout a global phage gene pool. And we have barely dipped our toes into that global pool.

Zhao Y, Wang K, Ackermann HW, Halden RU, Jiao N, & Chen F (2010). Searching for a "hidden" prophage in a marine bacterium. Applied and environmental microbiology, 76 (2), 589-95 PMID: 19948862

Five Questions About Lysogeny

by Merry

Lysogeny—a nasty time bomb or a mutually beneficial symbiosis? A prophage gone lytic will murder its host, but a symbiotic picture can well be argued. Here are some thoughts about the ongoing give-and-take. More details are still emerging.

1. If you are a phage, why be temperate?

A phage at the crossroads. Lambda (λ), the intensely-
studied temperate phage of E. coli, chooses between
the lytic pathway and lysogeny. Source.

For a phage, temperance offers the obvious advantage of providing a safe haven when host cells are few and far between or when conditions are not good for their rapid growth. Indeed, one does find more lysogens in nutrient-poor environments or during winter months. While nestled within a host chromosome, the prophage is faithfully replicated pari passu with host DNA. If the host clone prospers, so does the prophage. Here the prophage is protected from the heat-labile factors (proteases secreted by bacteria?) that can chew up a virion. Even while inside an intact virion, the phage DNA can be damaged by UV light. In sunlit waters, the number of infective virions is typically less than the number of intact virions, the difference being accounted for by virions that contain UV-damaged DNA. Prophages are also subject to UV damage, but being a segment of the host chromosome the damage is often mended by the host's DNA repair machinery. Of course, there are trade-offs here. Lysogeny eliminates the risky business of extra-cellular survival and locating a host, but one hungry protist can end the game for all concerned.

2. If you are a bacterium, why tolerate a prophage?

Source.

Having a prophage on board burdens you with more DNA to be replicated and with a passenger that just might kill you should conditions change. But said prophage protects you from infection by related phages and often supplies genes of immediate usefulness. Good examples of the latter are prophage-encoded toxins and other virulence factors, such as those essential for the pathogenesis of V. cholerae, E. coli O157, and C. diphtheriae. Even the temperate lambdoid phages of E. coli provide genes that make their hosts resistant to killing by serum complement. Some adaptations of nonpathogenic bacteria to a specific ecological niche almost surely also rely on useful prophage genes. It is thought that most of these genes had been acquired from previous hosts. Their conveyance by phage is central to horizontal gene transfer among prokaryotes.

This role of prophage as a source of new genes is of colossal importance. Bacterial genomes are composed of a core genome shared by virtually all members of a species, plus a halo of strain-specific genes. Prophage (along with related genomic islands) appear to be a primary source of these defining genes.

3. How do prophage genomes evolve?

Of Archaeal Periplasm & Iconoclasm

by Elio

Rough work, iconoclasm, but the only way to get at truth.
            Oliver Wendell Holmes

A thin section of Ignicoccus hospitalis under
the EM. C = cytoplasm. CM = "cytoplasmic"
membrane. P = periplasm. OS = outer sheath
(membrane). Bar = 0.5 µm. Source.

Biology is the iconoclast’s paradise. Over and over, cherished beliefs, some dating back for centuries, fall to the ground as exceptions to the rule are discovered. To the long list of such exceptions, we now add the finding by groups in Regensburg and Frankfurt that the outer membrane of an archaeon, Ignicoccus hospitalis, is energized and capable of generating ATP. Granted, this is a hyperthermophile who helped shatter the ancient belief that life at high temperatures is not possible, thus hardly a conformist. But this discovery is, to say the least, unexpected.

The old tenet is that the energetic business-end of prokaryotes is the cell membrane, whether surrounded by an outer membrane, as in Gram negatives, or not, as in Gram positives. I should know, I taught this for umpteen years. True, in Gram-negatives, energy can be transmitted to the outer membrane via the Ton system (a system that provides energy for the transport of iron siderophores, vitamin B12, and some colicins), but the reverse, making energy on the outer membrane and sending it to the cytoplasm, is not part of the old belief. Yet it’s been known for some time that a goodly number of bacteria can energize their outer membrane. They do this by having cytochromes inserted in the outer membrane where they carry out a process known as extracellular electron transfer. This ability stands the organisms in good stead, allowing them to utilize metals in rocks as electron acceptors.

Localization of A₁A_O ATP synthase on I. hospitalis cells in ultrathin sections. (A) Labeling
with antibodies against the purified ATPase complex, (B) Labeling with antibodies against the
membrane-bound subunit a. For both images the secondary antibody with ultrasmall gold
particles is visualized. C = cytoplasm. IM = inner membrane. V = vesicles in the periplasm.
OM = outer membrane. Bar = 1 µm. Source.

First of all, dual membrane systems have not be found in archaea other than Ignicoccus. What are the new conclusions about power generation in its outer membrane based on? Mainly on immunoelectron microscopy of sections using gold-labeled antibodies and immunofluorescence, which revealed that ATP synthase and H2:sulfur oxidoreductase are located entirely in the outer membrane. These two enzymes are required for energizing membranes and for ATP production. Thus, ATP can be expected be made in the outer membrane and released into the periplasm (which in this organism is huge—larger than the cytoplasm). You may ask, are these two enzymes also found in the inner membrane? The answer is no. Since the periplasm is so large and the two membranes so far apart, enzyme localization to one membrane or the other can be readily discerned. This introduces the question: which is the cytoplasmic membrane in this organism? The figure tells you something about the rare structural complexity of this organism. Note that the two membrane system is different from that of ordinary Gram-negatives, as here the outer membrane is not known to contain LPS or porins.

Co-culture of Ignicoccus spec. (green) and
Nanoarchaeum equitans (red). ss rRNA sequence-
specific fluorescence staining. Bar = 1 µm. Source.

Is the Ignicoccus story relevant to other prokaryotes? Who’s to say at this point. Ignicoccus is mightily idiosyncratic, e.g., it’s unique among archaea in having a two-membrane system. Not only does it grow at very high temperature and use reduction of elemental sulfur as its main energy source, but it also lives in intimate association with another archaeon, the smaller Nanoarchaeum equitans, which has a reduced genome and apparently gets its energy from its larger partner. The unusual ignicoccal ability to make ATP within its periplasm may help it to supply ATP to its associates across the outer membrane.

The authors propose a tantalizing notion: if the eukaryotic cell arose by an archaeon having swallowed a bacterium (hold on, we’re not getting into that discussion right now), then Ignicoccus or something like it would have been the ideal ancestor, able as it appears to be to donate ATP to anyone residing within its boundaries. True or not, one should further respect the outliers in the biological scheme of things as potential sources of novel and deeper relationships.

Research Blogging Citation
Kuper, U., Meyer, C., Muller, V., Rachel, R., & Huber, H. (2010). Energized outer membrane and spatial separation of metabolic processes in the hyperthermophilic Archaeon Ignicoccus hospitalis Proceedings of the National Academy of Sciences DOI: 10.1073/pnas.0911711107

Naegleria’s Split Morphology Disorder

by Psi Wavefunction

The complex structure of flagellates. EM section through
a mature flagellate Tetramitus. K1 = kinetosome. Cm =
cytostomal canal. FV = food vacuole. Rz = rhizoplast.
Source.

By default, a membrane-bound entity like a cell should be a spherical, formless blob. However, most cells are not such formless blobs, but rather have adopted one or more forms from a vast repertory of stunningly complex morphologies. To wit, see (and admire) the radiolarians, metazoan neurons, giant parabasalians or the endlessly weird and sophisticated ciliates. Even prokaryotes have a highly complex cellular structure, and are not, as some biochemists are prone to think, mere bags of enzymes. The deeper you venture into the realm of cellular diversity, the more awe-inspiring becomes the cornucopia of cellular structural and morphological variety. Luckily, there is some order to it as there are two fundamental 'genres' of cellular morphology, at least in the protists: flagellates and amoebae. Of course, there are also cysts, but since those are mostly resting stages (being a round ball isn't particularly helpful while feeding or fleeing from predators...) they can be ignored for now.

Cell shape depends on the cytoskeleton. As you know, its two main component systems are actin and tubulin, ignoring the plethora of miscellaneous proteins that are used for various structural jobs. Tubulin makes microtubules, the spindle fibers of mitosis, but is also important for the flagellar apparatus (we've yet to find one composed of actin and probably for good reasons). You also know that actin is a key player in cell motility and morphology in animal systems. It is also heavily involved in endomembrane trafficking within a cell, as well as endo- and exocytosis. If interested, a recent issue of Science has a nice overview of actin in morphogenesis and cell movement.

The role of the cytoskeleton in morphogenesis is much less clearly defined. It depends largely on the species. Plants, for example, rely very heavily on tubulin for morphology, with actin being a minor player. Amoeboid cells are primarily actin-based. In fact, amoeboid cells resort to tubulin largely for spindle formation during mitosis. They hate tubulin about as much as plants hate actin. Actin-based cells don't have to be amorphous; they are still able to achieve complex morphologies. But there is a positive correlation between amoeboid-ness and actin-ness ('actinity'?).

In contrast, flagellates are primarily tubulin-based. Of course, they still use actin for some intracellular work, but the shape depends largely on the whims of their microtubules. Perhaps not relying much on flagella allows the amoeboids to dispense with the microtubule organization pathways, thereby switching to actin. Flagellates, relying heavily on intact tubulin systems, may be less prone to losing their structure. Also, if you're a flagellate, you need shape for a modicum of streamlining. Try swimming around as a formless or floppy blob of some sort! Keep in mind that life at that scale is very different. Viscosity calls the shots when considering unicellular motility. Perhaps being hydrodynamic isn't even as important as simply retaining shape. Otherwise you'd be like a blob of molasses trying to swim through a sea of maple syrup. Not gonna get very far.

Whatever the reason, amoeboid cells tend to have a predominantly actin-based cytoskeleton, flagellates have a penchant for tubulin. Of course, not all organisms are decisive enough to make this commitment, so we've got amoeboflagellates in the middle.

Plenty of other organisms fancy transitioning between being more amoeboid or more flagellate. But few cells actually dispense with flagella and basal bodies altogether, to form them anew when special conditions arise. It's time to introduce one that does.