RNA

Essential or enduring understandings are often defined as the underlying core concepts or “big ideas” we’d like our students to remember when much of the course content has been forgotten. The central dogma of molecular biology and how cellular information is stored, used, and conveyed is one of the essential understandings students should retain after a course or unit in molecular biology or genetics. An additional enduring understanding is the relationships between DNA sequence, RNA sequence, mRNA production and processing, and the resulting polypeptide/protein product. A final big idea in molecular biology is the relationship between DNA mutation and polypeptide change. To engage students in these essential understandings in a Genetics course, I have developed a hands-on activity to simulate VDJ recombination. Students use a foldable type activity to splice out regions of a mock kappa light chain gene to generate a DNA sequence for transcription and translation. Students fold the activity several different times in multiple ways to “recombine” and generate several different DNA sequences. They then are asked to construct the corresponding mRNA and polypeptide sequence of each “recombined” DNA sequence and reflect on the products in a write-to-learn activity.

The goal of our study was to track changes in student understanding of the central dogma of molecular biology before and after taking a genetics course. Concept maps require the ability to synthesize new information into existing knowledge frameworks, and so the hypothesis guiding this study was that student performance on concept maps reveals specific central dogma misconceptions gained, lost, and retained by students. Students in a genetics course completed pre- and posttest concept mapping tasks using terms related to the central dogma. Student maps increased in complexity and validity, indicating learning gains in both content and complexity of understanding. Changes in each of the 351 possible connections in the mapping task were tracked for each student. Our students did not retain much about the central dogma from their introductory biology courses, but they did move to more advanced levels of understanding by the end of the genetics course. The information they retained from their introductory courses focused on structural components (e.g., protein is made of amino acids) and not on overall mechanistic components (e.g., DNA comes before RNA, the ribosome makes protein). Students made the greatest gains in connections related to transcription, and they resolved the most prior misconceptions about translation. These concept-mapping tasks revealed that students are able to correct prior misconceptions about the central dogma during an intermediate-level genetics course. From these results, educators can design new classroom interventions to target those aspects of this foundational principle with which students have the most trouble.

Research published in the Journal of Bacteriology demonstrates that a system of small RNAs help regulate the genetic regions called polysaccharide utilization loci (PULs) in the human gut-residing Bacteroides genus. First author Yanlu Cao and senior author C. Jeffrey Smith found that a small antisense RNA (sRNA) gene, donS, regulates activity of the don (for deglycosylation of N-glycans) locus. These sRNAs help the bacterium respond to multiple nutrient sources and may act as a regulatory mechanism to quickly modify gene expression under quickly changing nutrient conditions. The research was a collaboration between East Carolina University in Greenville, NC, and the University of Wúrzburg in Wúrzburg, Germany.

Several distinct types of Ebola therapeutic products are under development—some more or less conventional antiviral agents, while others depend on more recently developed techniques to disrupt viral gene activity or use antibodies to bind the virus and trigger host immune responses.

The U.S. Department of Energy (DOE) Joint Genome Institute (JGI) was established in 1997 to consolidate the department's programs and resources in DNA sequencing, informatics, and technology development. Soon after, the University of California, which manages the Lawrence Berkeley National Laboratory, where JGI was first situated, leased lab and office space in nearby Walnut Creek to house JGI activities. The early focus for JGI was the Human Genome Project. After its scientists completed their sequencing of three human chromosomes, however, JGI broadened its mandate in 2004 to become a national user facility, which now boasts thousands of users worldwide.

The highly conserved Nus factors of bacteria were discovered as essential host proteins for the growth of temperate phage λ in Escherichia coli. Later, their essentiality and functions in transcription, translation, and, more recently, in DNA repair have been elucidated. Close involvement of these factors in various gene networks and circuits is also emerging from recent genomic studies. We have described a detailed overview of their biochemistry, structures, and various cellular functions, as well as their interactions with other macromolecules. Towards the end, we have envisaged different uncharted areas of studies with these factors, including their participation in pathogenicity.

This chapter talks about the symbiosis of microbes and mitochondria. Mutations, including deletions of DNA, happen constantly, so the unused genes have long since disappeared from the genomes of mitochondria that inhabit our cells. In fact, mutations are still raining down on the genomes of our mitochondria, and these continue throughout our lives, contributing to various diseases and disorders. These malfunctions are the main reason we even think about our mitochondria. Another reason the mitochondria can live with so few genes is that many of the ancient genes of the mitochondrial ancestor have changed addresses, moving from the chromosome of the mitochondrion to our own chromosomes but sending their working products back to the mitochondrial homeland to carry out needed work. Some symbiotic bacteria have fewer than 200 genes, and the symbionts we call organelles can have even fewer, as in our own mitochondria with their miserly 15. Buchnera uses its ancestral genes for making the amino acids tryptophan and leucine, which are nutrients that a host needs. But these genes have been copied many times on tiny extra chromosomes (plasmids), an arrangement that allows hyperproduction of the amino acid products. Both symbionts and domesticated breeds can evolve extreme features over short periods on an evolutionary time scale. These extremes are produced mostly by exaggerating some functions, rather than by acquiring anything truly novel. And just like our domesticated animals and plants, symbionts themselves can be large, typically much larger than their free-living wild relatives.

Bacterial genetics has been strongly influenced by the work of John Roth and his laboratory. Work in John’s laboratory has covered a large number of topics, beginning with the genetic regulation of the histidine (his) biosynthetic operon and extending to many other metabolic pathways and genetic processes. John became interested in bacterial genetics while an undergraduate at Harvard University. The regulatory mutations were scattered widely around the genome and affected functions involved in translation (e.g., histidyl-tRNA synthetase, tRNAHis, and tRNA modifying and processing enzymes). Interest in genetics of tRNA led John’s lab to work on a variety of informational suppressors, including recessive nonsense suppressors and many classes of frameshift suppressors in which altered tRNAs caused translation to shift reading phase. John became interested in chromosomal duplications during early studies on nonsense suppressor tRNAs. Suppressor mutations that alter an essential tRNA type are lethal unless they arise in one copy of a preexisting duplication of the tRNA gene. The genetic tools developed in John’s lab helped change the perspectives on the pulsating rhythms of chromosome organization. Other work led to ideas on the evolution of bacterial operons by horizontal transfer and origins of new genes by selective amplification. In addition to his research accomplishments, John is an enthusiastic, stimulating speaker and teacher.

In environments where growth can be fast and production of proteins and metabolites is rapid, the stable isotope probing (SIP) approach is at its best. Rapid incorporation of stable isotopes into DNA, RNA, protein, and/or metabolites will be used routinely in medical and dental research, perhaps drawing these fields far closer toward environmental microbiology than they have been in the past. Eukaryotes evolved in a sea of Bacteria and Archaea, and it would be astounding if there were not many interdependent metabolic interactions that describe the total organism as a eukaryotic/bacterial/archaeal conglomerate. This understanding will be one of the great accomplishments of the next decades and will be greatly enhanced by SIP technologies at nearly every level (DNA, RNA, protein, and metabolites). The differences between hydrogen transfer and electron transfer could be significant and offer some major challenges to the understanding of microbial ecology: challenges that may be solved in part via the application of SIP approaches and others that will require new ways of thinking and experimentation. Several papers have appeared recently on extracellular electron transport. It is clear from the work in several laboratories that microbes have mechanisms available to them to donate electrons directly to solid surfaces and to take electrons directly from solid surfaces raising the possibility that an energy realm previously not thought possible by most microbiologists could exist in sedimentary environments, utilizing various types of extracellular electron transport mechanisms to deliver energy in the form of electrons among energy sources, cells, and electron acceptors.

In recent years it has become clear that complex regulatory circuits control the initiation step of DNA replication by directing the assembly of a multicomponent molecular machine (the orisome) that separates DNA strands and loads replicative helicase at oriC, the unique chromosomal origin of replication. This chapter discusses recent efforts to understand the regulated protein-DNA interactions that are responsible for properly timed initiation of chromosome replication. It reviews information about newly identified nucleotide sequence features within Escherichia coli oriC and the new structural and biochemical attributes of the bacterial initiator protein DnaA. It also discusses the coordinated mechanisms that prevent improperly timed DNA replication. Identification of the genes that encoded the initiators came from studies on temperature-sensitive, conditional-lethal mutants of E. coli, in which two DNA replication-defective phenotypes, "immediate stop" mutants and "delayed stop" mutants, were identified. The kinetics of the delayed stop mutants suggested that the defective gene products were required specifically for the initiation step of DNA synthesis, and subsequently, two genes, dnaA and dnaC, were identified. The DnaA protein is the bacterial initiator, and in E. coli, the DnaC protein is required to load replicative helicase. Regulation of DnaA accessibility to oriC, the ordered assembly and disassembly of a multi-DnaA complex at oriC, and the means by which DnaA unwinds oriC remain important questions to be answered and the chapter discusses the current state of knowledge on these topics.