Hypomorphic Glycosyltransferase Alleles and Recoding at Contingency Loci Influence Glycan Microheterogeneity in the Protein Glycosylation System of Neisseria Species

ABSTRACTAs more bacterial protein glycosylation systems are identified and characterized, a central question that arises is, what governs the prevalence of particular glycans associated with them? In addition, accumulating evidence shows that bacterial protein glycans can be subject to the phenomenon of microheterogeneity, in which variant glycan structures are found at specific attachment sites of a given glycoprotein. Although factors underlying microheterogeneity in reconstituted expression systems have been identified and modeled, those impacting natural systems largely remain enigmatic. On the basis of a sensitive and specific glycan serotyping system, microheterogeneity has been reported for the broad-spectrum,O-linked protein glycosylation system in species within the genusNeisseria. To elucidate the mechanisms involved, a genetic approach was used to identify a hypomorphic allele ofpglA(encoding the PglA galactosyltransferase) as a significant contributor to simultaneous expression of multiple glycoforms. Moreover, this phenotype was mapped to a single amino acid polymorphism in PglA. Further analyses revealed that manypglAphase-off variants (containing out-of-frame configurations in simple nucleotide repeats within the open reading frame) were associated with disproportionally high levels of theN,N′-diacetylbacillosamine–Gal disaccharide glycoform generated by PglA. This phenotype is emblematic of nonstandard decoding involving programmed ribosomal frameshifting and/or programmed transcriptional realignment. Together, these findings provide new information regarding the mechanisms of neisserial protein glycan microheterogeneity and the anticipatory nature of contingency loci.

Lack of Consistent Short Sequence Repeat Polymorphisms in Genetically Homologous Colonizing and Invasive Candida albicans Strains

ABSTRACT Short sequence repeats (SSRs), potentially representing variable numbers of tandem repeat (VNTR) loci, were identified for the human-pathogenic yeast species Candida albicans by computerized DNA sequence scanning. The individual SSR regions were investigated in different clinical isolates of C. albicans. Most of the C. albicans SSRs were identified as genuine VNTRs. They appeared to be present in multiple allelic variants and were demonstrated to be diverse in length among nonrelated strains. As such, these loci provide adequate targets for the molecular typing ofC. albicans strains. VNTRs encountered in other microbial species sometimes participate in regulation of gene expression and function as molecular switches at the transcriptional or translational level. Interestingly, the VNTRs identified here often encode polyglutamine stretches and are frequently located within genes potentially involved in the regulation of transcription. DNA sequencing of these VNTRs demonstrated that the length variability was restricted to the CAA/CAG repeats encoding the polyglutamine stretches. For these reasons, paired C. albicans isolates of similar genotype, either found as noninvasive colonizers or encountered in an invasive state in the same individual, were studied with respect to potentially invasion-related alterations in the VNTR profiles. However, none of the VNTRs analyzed thus far varied systematically with the transition from colonization to invasion. In contrast to the situation described for some prokaryotic species, this finding suggests that VNTRs of C. albicans may not simply function as contingency loci related to straightforward on/off regulation of invasion-related gene expression.