Mycobacterium tuberculosis Universal Stress Protein Rv2623 Regulates Bacillary Growth by ATP-Binding: Requirement for Establishing Chronic Persistent Infection

Abstract

Tuberculous latency and reactivation play a significant role in the pathogenesis of tuberculosis, yet the mechanisms that regulate these processes remain unclear. The Mycobacterium tuberculosis universal stress protein (USP) homolog, rv2623, is among the most highly induced genes when the tubercle bacillus is subjected to hypoxia and nitrosative stress, conditions thought to promote latency. Induction of rv2623 also occurs when M. tuberculosis encounters conditions associated with growth arrest, such as the intracellular milieu of macrophages and in the lungs of mice with chronic tuberculosis. Therefore, we tested the hypothesis that Rv2623 regulates tuberculosis latency. We observed that an Rv2623-deficient mutant fails to establish chronic tuberculous infection in guinea pigs and mice, exhibiting a hypervirulence phenotype associated with increased bacterial burden and mortality. Consistent with this in vivo growth-regulatory role, constitutive overexpression of rv2623 attenuates mycobacterial growth in vitro. Biochemical analysis of purified Rv2623 suggested that this mycobacterial USP binds ATP, and the 2.9-Å-resolution crystal structure revealed that Rv2623 engages ATP in a novel nucleotide-binding pocket. Structure-guided mutagenesis yielded Rv2623 mutants with reduced ATP-binding capacity. Analysis of mycobacteria overexpressing these mutants revealed that the in vitro growth-inhibitory property of Rv2623 correlates with its ability to bind ATP. Together, the results indicate that i) M. tuberculosis Rv2623 regulates mycobacterial growth in vitro and in vivo, and ii) Rv2623 is required for the entry of the tubercle bacillus into the chronic phase of infection in the host; in addition, iii) Rv2623 binds ATP; and iv) the growth-regulatory attribute of this USP is dependent on its ATP-binding activity. We propose that Rv2623 may function as an ATP-dependent signaling intermediate in a pathway that promotes persistent infection.

Strains of the East Asian (W/Beijing) Lineage ofMycobacterium tuberculosisAre DosS/DosT-DosR Two-Component Regulatory System Natural Mutants

“…While their specific functions remain largely unknown, the Usp superfamily-like bacterial 2CS-are involved in stress resistance, adaptation to energy deficiency, cell motility, and adhesion (42,44). In M. tuberculosis, the Usp encoded by the Rv2623 gene is part of the DosR regulon and appears to regulate bacterial growth in an ATPdependent manner (14). There is also at least one example in the E. coli literature where a Usp specifically modulates the activity of a 2CS (25).…”

An obligately aerobic soil bacterium activates fermentative hydrogen production to survive reductive stress during hypoxia

“…Numerous components of the DosR regulon of M. smegmatis are also DosR dependent in pathogenic mycobacteria, including multiple universal stress proteins (30,31), a ribosome stabilizingfactor (32,33), and putative energy metabolism genes (e.g., diacylglycerol acyltransferases, nitroreductases) (34,35) (Tables S1 and S2). The DosR regulon of M. smegmatis also controls the expression of some genes that are not DosR dependent or are absent from the pathogen (Table S2), most notably 10 genes involved in hydrogen metabolism and 6 genes linked to sulfur metabolism ( Fig.…”

Genome-Wide Association Mapping Combined with Reverse Genetics Identifies New Effectors of Low Water Potential-Induced Proline Accumulation in Arabidopsis

“…The UspA domain kinase AT5G35380 was associated with the SNP of the second lowest P value in our analysis, and mutants of two additional UspA proteins (without the kinase domain) also affected Pro accumulation. In bacteria, UspA domain proteins control stress responses and growth through adenine nucleotide binding and may act as switches that detect cellular energy or metabolic status (Persson et al, 2007;Drumm et al, 2009). Arabidopsis has large families of both UspA kinases and UspA proteins without kinase domains (Kerk et al, 2003).…”