Transcription of the Streptococcus pyogenes Hyaluronic Acid Capsule Biosynthesis Operon Is Regulated by Previously Unknown Upstream Elements

Abstract

The important human pathogen Streptococcus pyogenes (group A Streptococcus [GAS]) produces a hyaluronic acid (HA) capsule that plays critical roles in immune evasion. Previous studies showed that the hasABC operon encoding the capsule biosynthesis enzymes is under the control of a single promoter, P1, which is negatively regulated by the two-component regulatory system CovR/S. In this work, we characterize the sequence upstream of P1 and identify a novel regulatory region controlling transcription of the capsule biosynthesis operon in the M1 serotype strain MGAS2221. This region consists of a promoter, P2, which initiates transcription of a novel small RNA, HasS, an intrinsic transcriptional terminator that inefficiently terminates HasS, permitting read-through transcription of hasABC, and a putative promoter which lies upstream of P2. Electrophoretic mobility shift assays, quantitative reverse transcription-PCR, and transcriptional reporter data identified CovR as a negative regulator of P2. We found that the P1 and P2 promoters are completely repressed by CovR, and capsule expression is regulated by the putative promoter upstream of P2. Deletion of hasS or of the terminator eliminates CovR-binding sequences, relieving repression and increasing read-through, hasA transcription, and capsule production. Sequence analysis of 44 GAS genomes revealed a high level of polymorphism in the HasS sequence region. Most of the HasS variations were located in the terminator sequences, suggesting that this region is under strong selective pressure. We discovered that the terminator deletion mutant is highly resistant to neutrophil-mediated killing and is significantly more virulent in a mouse model of GAS invasive disease than the wild-type strain. Together, these results are consistent with the naturally occurring mutations in this region modulating GAS virulence.

Impact of contusion injury on intramuscular emm1 group a streptococcus infection and lymphatic spread

“…5/5 mucoid isolates from spleen showed a single nucleotide polymorphism (SNP) in covR resulting in a base and amino acid change, ACC (threonine) to CCC (proline) occurring at codon 65 (out of 228). However, in 5/5 mucoid colonies obtained from the ipsilateral lymph node, a 213bp deletion in the promoter of the hasABC operon was detected, in a region incorporating the recently described P2 promoter region[25], hasS region, and one CovR DNA binding site (Figure 5(d)). Compared to the parental strain, no other SNPs were identified in either the colonies from the spleen or the lymph node.…”

“…To determine the basis for hyper-encapsulation, in one experiment, we undertook genome sequencing of representative isolates from lymph node and spleen from a single mouse. Although isolates from the spleen demonstrated a mutation in covR , which might be predicted from many other studies [9,32], the hyper-encapsulated GAS variants in the draining lymph node demonstrated a deletion in the hasABC promoter which leads to loss of one of the reported CovR binding sites[25], thereby de-repressing capsule production specifically [25]. The hyper-encapsulated promoter mutant strain (P2) had enhanced fitness with regard to transit to, and/or retention in, the draining lymph node consistent with the reported interaction between hyaluronan and the lymphatic receptor LYVE-1 [22].…”

“…Deletion of 213bp (indicated by red dotted line box) in hasABC promoter region identified in all five hyper-encapsulated colony variants from lymph node. Deleted region includes P2 promoter, regulator sRNA hasS region, and CovR binding site for the P1 promoter [25,34]. Equivalent position of deletion in completed reference genome MGAS5005 (NC_007297.2) is indicated as 1,818,451-1818663bp.…”

Multimerization of the Virulence-Enhancing Group A Streptococcus Transcription Factor RivR Is Required for Regulatory Activity

“…The parental strain used in this study was the serotype M1 GAS strain MGAS2221, which we and others have characterized extensively (23,35,36). A list of all GAS strains used in this study is presented in Table 1.…”

“…While not studied in all Mga-like family members, the Mga and AtxA regulators are able to self-dimerize via an N-terminal dimerization domain (21,22). For regulatory activity, Mga dimers come together to form higher-order oligomers, with the multimerization domain being located in the unstructured C-terminal region (23).…”