This lab at UCLA is doing some pretty neat stuff with very small microscopes. I read about them in the Feb 2013 National Geographic but the article was very vague on details. They're working on developing mobile devices that can diagnose diseases like malaria using microscopy. Very useful.
Today's paper is:
Pneumocystis jirovecii is a fungus that isn't usually pathogenic but is deadly to immunocompromised patients, most prominently those with AIDS. As with many other parasitic microbes colonizing human tissues, P. jirovecii can't be cultured in the lab. This disadvantage makes working with P. jirovecii nearly impossible if researchers seek to use genetic or molecular techniques. This looks like a case for sequencing! Wait, no, hold on. Researchers generally need to culture microbes to get enough genomic DNA for sequencing. The authors of this paper bypass that issue by collecting the fungus direct from the lungs of infected patients (they also used immunoprecipitation and random amplification, so this by no means an instance of single-cell sequencing).
Collecting samples of bronchoalveolar lavage fluid (or, if you prefer, the ridiculous-sounding acronym BALFs) provides plenty of fungus to work with. The samples are also rich in other genetic material from human cells, bacteria, viruses, and on occasion other species of Pneumocystis. The authors had to extract the P. jirovecii genome from the mess -- luckily, the genome of related species and rat pathogen P. carinii has already been sequenced and partially assembled. Process of elimination allowed for removal of the non-Pneumocystis sequences, then genome comparison and RNAseq helped finish and annotate the rest. More than just a de novo genome assembly using a messy metagenomic data source, this work is an example of how availability of massive quantities of genomic data make some projects entirely feasible.
It's a snowy, slushy mess out there! It's certainly been more wintry in Richmond before but rarely the kind of weather in which trees actively pelt me with slushballs on the walk to campus. It's still snowing, too. Danielle and I were planning on driving to Delaware this evening or tomorrow morning so I hope most of it gets cleaned up by then.
Today's paper is: Rao and Black - Structure and assembly of bacteriophage T4 head (2010) Virology Journal.
This is a review of the essential observations of the phage T4 head structure. Next to the famous lambda phage, T4 and its fellow T-even phages may be some of the most closely-studied viruses. T4 has that classic phage structure: an icosohedral head, a contractile tail, and a baseplate at the end of the tail, plus six spindly fibers on the baseplate. Cryo-EM has helped to define the structures of the primary T4 head proteins (gp23 and gp24) down to about 0.3 nm or 3 angstrom, which is a resolution that's hard to improve on. The gp23 protein demonstrates the classical HK97 fold and relies upon the E. coli chaperone GroEL and a phage-encoded cochaperone gp31 to reach its final form.
The paper digresses a bit to discuss phage display. I wasn't aware of how this technique was being used in recent years, but antigens fused to T4 capsids have been shown to be effective vaccines against foot and mouth disease virus, anthrax toxin and potentially even tumor metastasis. The authors aren't clear about how phage help with vaccine delivery. I suspect that they may have improved long-term storage capabilities when compared to antigen alone but the overall number of fusion proteins is limited by copy number per phage capsid; it can't really exceed the 870 copies of Soc already bound to the capsid and may also be limited by steric hindrance. Displaying antigens on phage rather than on their own may increase the immune response but I'm curious if it makes sense to use phage instead of a chemical adjuvant. The folks who did the anti-tumor study cite some references claiming as much and also claim that phage have the benefit of causing few side effects. I'd still call potential disruption of enteric bacteria a notable side effect.
Other interesting notes about the T4 capsid:
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| The T4 head structure. Looks kinda like a pineapple. |
The paper digresses a bit to discuss phage display. I wasn't aware of how this technique was being used in recent years, but antigens fused to T4 capsids have been shown to be effective vaccines against foot and mouth disease virus, anthrax toxin and potentially even tumor metastasis. The authors aren't clear about how phage help with vaccine delivery. I suspect that they may have improved long-term storage capabilities when compared to antigen alone but the overall number of fusion proteins is limited by copy number per phage capsid; it can't really exceed the 870 copies of Soc already bound to the capsid and may also be limited by steric hindrance. Displaying antigens on phage rather than on their own may increase the immune response but I'm curious if it makes sense to use phage instead of a chemical adjuvant. The folks who did the anti-tumor study cite some references claiming as much and also claim that phage have the benefit of causing few side effects. I'd still call potential disruption of enteric bacteria a notable side effect.
Other interesting notes about the T4 capsid:
- The T4 packaging motor packages DNA at about 2 kb/s. It's an ATP-dependent process - no silly physics tricks here, though it's incredibly efficient (the authors state that this motor is "...approximately twice as powerful as a typical automobile engine", whatever they mean by 'typical'). The T4 genome is 171 kb so the motor gets the whole thing and then some packaged in less than 5 minutes. The authors must have summarized some numbers as a constant rate of 2000 kb/s should get it all packaged in less than 2 minutes, so the rest of the time may proceed more slowly or may be subject to something like strand slippage.
- The structure of the T4 portal complex is conserved across many phages and in HSV, which is really just an overgrown phage anyway. Studies really don't agree about how it works and I'm not inclined to bet on any one model at the moment.