The Hsp70-like StkA functions between T4P and Dif signaling proteins as a negative regulator of exopolysaccharide inMyxococcus xanthus

Abstract

Myxococcus xanthus displays a form of surface motility known as social (S) gliding. It is mediated by the type IV pilus (T4P) and requires the exopolysaccharide (EPS) to function. It is clear that T4P retraction powers S motility. EPS on a neighboring cell or deposited on a gliding surface is proposed to anchor the distal end of a pilus and trigger T4P retraction at its proximal end. Inversely, T4P has been shown to regulate EPS production upstream of the Dif signaling pathway. Here we describe the isolation of two Tn insertions at the stk locus which had been known to play roles in cellular cohesion and formation of cell groups. An insertion in stkA (MXAN_3474) was identified based on its ability to restore EPS to a pilA deletion mutant. The stkA encodes a DnaK or Hsp70 homolog and it is upstream of stkB (MXAN_3475) and stkC (MXAN_3476). A stkB insertion was identified in a separate genetic screen because it eliminated EPS production of an EPS+ parental strain. Our results with in-frame deletions of these three stk genes indicated that the stkA mutant produced increased level of EPS while stkB and stkC mutants produced less EPS relative to the wild type. S motility and developmental aggregation were affected by deletions of stkA and stkB but only minimally by the deletion of stkC. Genetic epistasis indicated that StkA functions downstream of T4P but upstream of the Dif proteins as a negative regulator of EPS production in M. xanthus.

sRNA‐pathway genes regulating myxobacterial development exhibit clade‐specific evolution

“…Such processing of a primary transcript to an active non-coding regulatory small RNA is reminiscent of similar mechanisms in the generation of eukaryotic miRNAs (Shukla et al, 2011). In M. xanthus, StkA functions as a negative regulator of extracellular polysaccharide production (Moak, Black, Wallace, Li, & Yang, 2015) and, based on our findings here, might promote the function of a protein (or proteins) necessary for Pxr production, stability or function during development. In M. xanthus, StkA functions as a negative regulator of extracellular polysaccharide production (Moak, Black, Wallace, Li, & Yang, 2015) and, based on our findings here, might promote the function of a protein (or proteins) necessary for Pxr production, stability or function during development.…”

“…The stkA locus encodes a protein homologous to the chaperone DnaK and the Hsp70 family of proteins that facilitate the folding of newly synthesized proteins and the refolding of proteins denatured or misfolded by heat shock in many organisms (Genevaux et al, 2007). In M. xanthus, StkA functions as a negative regulator of extracellular polysaccharide production (Moak, Black, Wallace, Li, & Yang, 2015) and, based on our findings here, might promote the function of a protein (or proteins) necessary for Pxr production, stability or function during development.…”

The type IV pilus assembly ATPase PilB functions as a signaling protein to regulate exopolysaccharide production in Myxococcus xanthus

“…Genetic mapping by transformation using genomic DNA 39 , 41 determined that two out of the five isolates likely had suppressor mutations in the mutagenized pilBTC genes as they are linked to the kanamycin resistant (Kan R ) marker carried by the integrative plasmid. The other three, which must have occurred elsewhere 29 , 36 , 39 , 41 , 42 , were not pursued further in this study.…”

“…Since mutations in dif are epistatic to those in T4P or pil genes, T4P has been proposed to function as a sensory apparatus that perceives and transmits signals to the Dif proteins downstream. The communication between T4P and Dif is mediated in part by the negative regulator StkA, a DnaK-like protein 34 , 35 that acts downstream of T4P but upstream of Dif 36 . Many questions remain concerning the mechanism of EPS regulation by this pathway although the transcription of eps genes does not appear to be the target of this regulation 37 .…”

Conditional requirement of SglT for type IV pili function and S-motility in Myxococcus xanthus

“…In accordance with the qualitative assays, WT and SglT had equivalent EPS levels, 15.0 and 15.9 % of Trypan blue bound, respectively. As a comparison, the Dsp (EPS – ) mutant bound only 0.6 %, while the Stk (‘sticky’) mutant, which overproduces EPS [41], bound 21.0 % of the dye ().…”