Abstract
Regulated RNA turnover is vital for the control of gene expression in all cellular life. In Escherichia coli, this process is largely controlled by a stable degradosome complex containing RNase E and a variety of additional enzymes. In the Firmicutes phylum, species lack RNase E and often encode the paralogous enzymes RNase J1 and RNase J2. Unlike RNase J1, surprisingly little is known about the regulatory function and protein interactions of RNase J2, despite being a central pleiotropic regulator for the streptococci and other closely related organisms. Using crosslink coimmunoprecipitation in Streptococcus mutans, we have identified the major proteins found within RNase J2 protein complexes located in the cytoplasm and at the cell membrane. In both subcellular fractions, RNase J2 exhibited the most robust interactions with RNase J1, while additional transient and/or weaker “degradosome-like” interactions were also detected. In addition, RNase J2 exhibits multiple novel interactions that have not been previously reported for any RNase J proteins, some of which were highly biased for either the cytoplasmic or membrane fractions. We also determined that the RNase J2 C-terminal domain (CTD) encodes a structure that is likely conserved among RNase J enzymes and may have an analogous function to the C-terminal portion of RNase E. While we did observe a number of parallels between the RNase J2 interactome and the E. coli degradosome paradigm, our results suggest that S. mutans degradosomes are either unlikely to exist or are quite distinct from those of E. coli.
Highlights
RNase activity is an essential component of bulk RNA turnover, RNA processing, and posttranscriptional gene regulation
The composition of the E. coli degradosome has been well characterized and consists of a mixture of enzymes directly involved in RNA metabolism, such as the DEAD-box RNA helicase (RhlB) (Miczak et al, 1996; Py et al, 1996), polynucleotide phosphorylase (PNPase) (Miczak et al, 1996; Liou et al, 2001), polyphosphate kinase (PPK) (Blum et al, 1997), and RNase II (Lu and Taghbalout, 2014)
Degradosome-like protein complexes have been detected with RNase Y and include interactions with RNases J1/J2, PNPase, DEAD-box RNA helicase (CshA), and the glycolytic enzymes enolase (Eno) and phosphofructokinase (PfkA) (Lehnik-Habrink et al, 2010, 2011)
Summary
RNase activity is an essential component of bulk RNA turnover, RNA processing, and posttranscriptional gene regulation. Degradosome-like protein complexes have been detected with RNase Y and include interactions with RNases J1/J2, PNPase, DEAD-box RNA helicase (CshA), and the glycolytic enzymes enolase (Eno) and phosphofructokinase (PfkA) (Lehnik-Habrink et al, 2010, 2011). While multiple RNA metabolizing enzymes interact with RNase Y in B. subtilis (Commichau et al, 2009), it is not yet clear whether these interactions mediate stable membrane localization comparable to the role of RNase E (Khemici et al, 2008; Gorna et al, 2012). Both the N- and C-terminal domains of RNase J2 are able to localize to the cell membrane, while the CTD serves as a site of multiple protein–protein interactions
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