SCDFinder, a web-based tool for the identification of putative novel ATM and ATR targets.

MOTIVATION: The S/TQ cluster domain (SCD) constitutes a new type of protein domain that is not defined by sequence similarity but by the presence of multiple S/TQ motifs within a variable stretch of amino acids. SCDs are recognized targets for DNA damage response (DDR) kinases like ATM and ATR. Characterizing DDR targets is of significant interest. The aim of this work was to develop a web-based tool to allow for easy identification and visualization of SCDs within specific proteins or in whole proteome sets, a feature not supported by current domain and motif search tools. RESULTS: We have developed an algorithm that (i) generates a list of all proteins in an organism containing at least one user-defined SCD within their sequence, or (ii) identifies and renders a visual representation of all user-defined SCDs present in a single sequence or batch of sequences. AVAILABILITY AND IMPLEMENTATION: The application was developed using Pearl and Python, and is available at the following URL: http://ustbioinfo.webfactional.com/scd/.

Experimental evolution of an essential Bacillus gene in an E. coli host.

The acquisition of foreign genes by HGT potentially greatly speeds up adaptation by allowing faster evolution of beneficial traits. The evolutionary integration of novel genes into host gene expression and physiology is critical for adaptation by HGT, but remains largely unknown. We are exploring the evolutionary consequences of gene acquisition in populations of Escherichia coli in real time. A plasmid bearing the genes necessary for sucrose catabolism was constructed and introduced into a single E. coli genotype. Wild-type E. coli is generally incapable of utilizing sucrose, but E. coli transformants were able to grow on sucrose as a sole carbon and energy source, albeit poorly. Twelve replicate populations were initiated and propagated in sucrose minimal media for 300 generations. Over this time, we observed large fitness improvements in the selected environment. These results demonstrate the potential for HGT to substantially increase microbial niche breadth.

Biohazard potential of putative Martian organisms during missions to Mars.

Exploration Class missions to Mars will require precautions against potential contamination by any native microorganisms that may be incidentally pathogenic to humans. While the results of NASA's Viking biology experiments of the 1970s have been generally interpreted as inconclusive for surface organisms, and attributed to active but nonbiological chemistries, the possibility of native surface life has never been ruled out completely. It is possible that, prior to the first human landing on Mars, robotic craft and sample return missions will provide enough data to know with certainty whether future human landing sites harbor extant life forms. If native life were found to exist, it would be problematic to determine whether any of its species might present a medical danger to astronauts. Therefore, it will become necessary to assess empirically the risk that the planet contains pathogens based on terrestrial examples of pathogenicity and to take a reasonably cautious approach to biohazard protection. A survey of terrestrial pathogens was conducted with special emphasis on those whose evolution has not depended on the presence of animal hosts. The history of the development and implementation of Apollo anti-contamination protocol and recommendations of the National Research Council's Space Studies Board regarding Mars were reviewed. Organisms can emerge in Nature in the absence of indigenous animal hosts and both infectious and non-infectious human pathogens are therefore theoretically possible on Mars. Although remote, the prospect of Martian surface life, together with the existence of a diversity of routes by which pathogenicity has emerged on Earth, suggests that the probability of human pathogens on Mars, while low, is not zero. Still, since the discovery and study of Martian life can have long-term benefits for humanity, the risk that Martian life might include pathogens should not be an obstacle to human exploration. As a precaution, it is recommended that EVA (extravehicular activity) suits be decontaminated when astronauts enter surface habitats upon returning from field activity and that biosafety protocols approximating laboratory BSL 2 be developed for astronauts working in laboratories on the Martian surface. Quarantine of astronauts and Martian materials arriving on Earth should also be part of a human mission to Mars, and this and the surface biosafety program should be integral to human expeditions from the earliest stages of the mission planning.

16S rRNA beacons for bacterial monitoring during human space missions.

Microorganisms are unavoidable in space environments and their presence has, at times, been a source of problems. Concerns about disease during human space missions are particularly important considering the significant changes the immune system incurs during spaceflight and the history of microbial contamination aboard the Mir space station. Additionally, these contaminants may have adverse effects on instrumentation and life-support systems. A sensitive, highly specific system to detect, characterize, and monitor these microbial populations is essential. Herein we describe a monitoring approach that uses 16S rRNA targeted molecular beacons to successfully detect several specific bacterial groupings. This methodology will greatly simplify in-flight monitoring by minimizing sample handling and processing. We also address and provide solutions to target accessibility problems encountered in hybridizations that target 16S rRNA.

NCIR: a database of non-canonical interactions in known RNA structures.

The secondary and tertiary structure of an RNA molecule typically includes a number of non-canonical base-base interactions. The known occurrences of these interactions are tabulated in the NCIR database, which can be accessed from http://prion.bchs.uh.edu/bp_type/. The number of examples is now over 1400, which is an increase of >700% since the database was first published. This dramatic increase reflects the addition of data from the recently published crystal structures of the 50S (2.4 A) and 30S (3.0 A) ribosomal subunits. In addition, non-canonical interactions observed in published crystal and NMR structures of tRNAs, group I introns, ribozymes, RNA aptamers and synthetic oligonucleotides are included. Properties associated with these interactions, such as sequence context, sugar pucker conformation, glycosidic angle conformation, melting temperature, chemical shift and free energy, are also reported when available. Out of the 29 anticipated pairs with at least two hydrogen bonds, 28 have been observed to date. In addition, several novel examples, not generally predicted, have also been encountered, bringing the total of such pairs to 36. Added to this list are a variety of single, bifurcated, triple and quadruple interactions. The most common non-canonical pairs are the sheared GA, GA imino, AU reverse Hoogsteen, and the GU and AC wobble pairs. The most frequent triple interaction connects N3 of an A with the amino of a G that is also involved in a standard Watson-Crick pair.

The ATM- and ATR-related SCD domain is over-represented in proteins involved in nervous system development.

ATM and ATR are cellular kinases with a well-characterized role in the DNA-damage response. Although the complete set of ATM/ATR targets is unknown, they often contain clusters of S/TQ motifs that constitute an SCD domain. In this study, we identified putative ATM/ATR targets that have a conserved SCD domain across vertebrates. Using this approach, we have identified novel putative ATM/ATR targets in pathways known to be under direct control of these kinases. Our analysis has also unveiled significant enrichment of SCD-containing proteins in cellular pathways, such as vesicle trafficking and actin cytoskeleton, where a regulating role for ATM/ATR is either unknown or poorly understood, hinting at a much broader and overarching role for these kinases in the cell. Of particular note is the overrepresentation of conserved SCD-containing proteins involved in pathways related to neural development. This finding suggests that ATM/ATR could be directly involved in controlling this process, which may be linked to the adverse neurological effects observed in patients with mutations in ATM.

Small RNA sequences are readily stabilized by inclusion in a carrier rRNA.

This laboratory previously showed that an RNA derived from 5S ribosomal RNA could be used as a carrier to harbor a nucleic acid "tag" for monitoring genetically engineered or naturally occurring bacteria. The prototype system expressed a specific tagged RNA that was stable and accumulated to high levels. For such a system to be useful there should, however, be little limitation on the sequence composition and length of the insert. To test these limitations, a collection of insertion sequences were created and introduced into the artificial 5S rRNA cassette. This library consisted of random 13- and 50-base oligonucleotides that were inserted into the carrier RNA. We report here that essentially all of the insert-containing RNAs are stable and accumulate to detectable levels. Tagged RNAs were produced by both plasmid-borne and chromosomally integrated expression systems in E. coli and several Pseudomonas strains without obvious effect on the host cell. It is anticipated that in addition to its intended use in environmental monitoring, this system can be used for in vivo selection of useful artificial RNAs. Because the carrier lends stability to the RNAs, the system may also be useful in RNA production.

Implementation of a Service-learning Module in Medical Microbiology and Cell Biology Classes at an Undergraduate Liberal Arts University.

Here we discuss the implementation of a service-learning module in two upper-division biology classes, Medical Microbiology and Cell Biology. This exciting hands-on learning experience provided our students with an opportunity to extend their learning of in-class topics to a real-life scenario. Students were required to volunteer their time (a minimum of 10 hours in a semester) at an under-served clinic in Houston, Texas. As they interacted with the personnel at the clinic, they were asked to identify the most prevalent disease (infectious for Medical Microbiology, and cellular-based for Cell) seen at the clinic and, working in groups, come up with educational material in the form of a display or brochure to be distributed to patients. The material was meant to educate patients about the disease in general terms, as well as how to recognize (symptoms), prevent and treat it. Students were required to keep a reflective journal in the form of a blog throughout the semester, and present their final materials to the class orally. Students were surveyed about their opinion of the experience at the end of the semester. The vast majority of student participants felt that the project was a positive experience and that it helped them develop additional skills beyond what they learn in the classroom and understand how lecture topics relate to every day life.

Ribosome origins: the relative age of 23S rRNA Domains.

The modern ribosome and its component RNAs are quite large and it is likely that at an earlier time they were much smaller. Hence, not all regions of the modern ribosomal RNAs (rRNA) are likely to be equally old. In the work described here, it is hypothesized that the oldest regions of the RNAs will usually be highly integrated into the machinery. When this is the case, an examination of the interconnectivity between local RNA regions can provide insight to the relative age of the various regions. Herein, we describe an analysis of all known long-range RNA/RNA interactions within the 23S rRNA and between the 23S rRNA and the 16S rRNA in order to assess the interconnectivity between the usual Domains as defined by secondary structure. Domain V, which contains the peptidyl transferase center is centrally located, extensively connected, and therefore likely to be the oldest region. Domain IV and Domain II are extensively interconnected with both themselves and Domain V. A portion of Domain IV is also extensively connected with the 30S subunit and hence Domain IV may be older than Domain II. These results are consistent with other evidence relating to the relative age of RNA regions. Although the relative time of addition of the GTPase center can not be reliably deduced it is pointed out that the development of this may have dramatically affected the progenotes that preceded the last common ancestor.