title: Cellular localization of RNA degradation and processing components in Escherichia coli.
abstract: The ability to study the localization and organization of proteins within the tiny cells of bacteria, such as Escherichia coli, has paved the way for a new and exciting era of prokaryotic cellular biology. Previously unrecognized levels of spatiotemporal and supramolecular organization of proteins have been revealed within the prokaryotic cell that had long been assumed as a "bag of enzymes." Immunofluorescence (IF) microscopy, which involves cellular immunostaining of native proteins with fluorescently labeled antibodies, is relatively laborious and requires cell fixation and highly specific antibodies. However, IF microscopy allows localization studies of native proteins expressed to their normal cellular levels, as opposed to labeling proteins with large fluorescent tag that can alter protein abundance dependent on changes in mRNAs and/or proteins stability, or whose detection can require overexpression of labeled proteins. In addition, when antibodies against native proteins are not available or lack specificity, epitope tags such as hemagglutinin (HA) or Flag can be used to label chromosomally expressed proteins. The short Flag- and HA-tag, eight or nine amino acids, are unlikely to interfere with the localization or function of the proteins. We describe and discuss here the use of fluorescence microscopy for determination of cellular organization of protein components of the E. coli RNA processing and degradation machinery. We present examples of cellular organization patterns visualized by light microscopy, either by IF microscopy of native and epitope-tagged proteins in fixed cells, or by fluorescence labeling of the proteins in live cells.
authors.affiliation: Univ. Paris Diderot, Sorbonne Paris Cité, Paris, 75013, France.
authors.forename: Véronique
authors.initials: V
authors.lastname: Arluison
authors.forename: Aziz
authors.initials: A
authors.lastname: Taghbalout
mesh_terms: D004926:Escherichia coli / Q000378:metabolism*; D029968:Escherichia coli Proteins / Q000378:metabolism; D012329:RNA, Bacterial / Q000378:metabolism*
chemical_list: D029968:Escherichia coli Proteins; D012329:RNA, Bacterial