WalK（S221P）mutation promotes production of Staphylococcus aureus capsule

To explore the role and possible mechanism of WalK（S221P）mutation in promoting capsule production of Staphylococcus aureus（S. aureus）.

RNA sequencing（RNA-seq）was performed to detect the effect of WalK（S221P）mutation on gene expression of S. aureus strain XN108. RT-qPCR was used to verify the capsule gene variation caused by WalK（S221P）mutation. Capsule staining and transmission electron microscopy（TEM）were employed to determine capsule biosynthesis of S. aureus. The possible regulatory genes involved in WalK（S221P）regulation were screened by using RT-qPCR. The binding site of WalKR on the regulatory regions of mgrA gene was predicted, and electrophoresis mobility shift assay（EMSA）was conducted to detect the direct binding activity of WalK-activated WalR on the mgrA DNA probe.

RNA-seq revealed that the expression of 196 genes altered after WalK（S221P）mutation in S. aureus XN108, and among these genes, 16 genes involved in capsule biosynthesis were upregulated remarkably. RT-qPCR confirmed the upregulation of capA, capH, and capK in the WalK（S221P）-carried S. aureus strains, including XN108 and K-Newman. Capsule staining showed that XN108 had more capsule production than its counterpart XN108-R. TEM observation demonstrated that XN108 had thicker capsule than XN108-R. RT-qPCR indicated an increasing expression of mgrA gene in the WalK（S221P）-carried XN108 when compared with that in XN108-R. A putative binding site of WalR on the regulatory regions of mgrA gene was predicted, and the direct binding activity of WalK-activated WalR on the mgrA regulatory fragment was verified by EMSA.

WalK（S221P）promotes the production of S. aureus capsule, and the underlying mechanism might be associated with the upregulation of MgrA, a positive regulator for S. aureus capsule biosynthesis.