Single-molecule analysis of transcription-elongation regulation mechanisms in living bacteria

Our study uses ultra-sensitive microscopes to observe important processes occurring during gene expression, the path that converts genetic information stored in DNA (present in cells in the form of chromosomes) to the manufacturing of proteins and other molecules that serve as the machinery, sensors, and structural framework of living cells.

Specifically, the work focuses on the process of gene transcription, which is performed by protein machines called RNA polymerases, which read DNA and copy the information into RNA molecules. RNA can serve either as a message (i.e., messenger RNA or "mRNA") or become part of other large machinery, such as ribosomal RNA or "rRNA", which is part of the ribosomes, the machines that make proteins in the cell. Transcription is further controlled by proteins known as transcriptional regulators, ensuring that the right genes are expressed at the right time, the right place, and at the required level.

In particular, we are studying transcriptional regulators NusG and RfaH, which couple RNA polymerase to other machineries (such as the ribosome) during the phase of transcription elongation, where the RNA polymerase rapidly extends RNA molecules. NusG and RfaH are very important regulators since the family they form controls transcription in all living organisms; RfaH is also a biomedically important, since it allows many pathogenic bacteria to turn on genes that can cause disease and help evade antibiotic treatments.

Much of what we know about how RNA polymerase and transcription regulators work comes from studies with purified proteins and DNA in the test tube; these involve simple mixtures of RNA polymerase with DNA sequences and transcription regulators that can accelerate or slow down transcription. However, the mechanisms of transcription in actual living organisms and cells can be very different, both due to the myriad of other biological components present in cells, and due to the way that genes are packaged in the "bacterial...