Elucidating essential pathways for the pathogenicity of Neisseria gonorrhoeae is crucial for the discovery of novel therapeutics required to combat the threat of extensively drug-resistant strains. Pathways of sulfur metabolism have been proposed as attractive targets as they are absent in mammals, and cysteine, the central node in sulfur metabolism, is critical to a broad range of cellular functions. Accordingly, the acquisition of sulfur sources to fulfil cysteine requirements is a crucial determinant of bacterial survival. However, in N. gonorrhoeae these pathways remain poorly defined.
Here, we investigate the metabolic flexibility of N. gonorrhoeae with respect to sulfur acquisition and examine how the gonococcus adapts to conditions of sulfur limitation. By integrating bioinformatic analyses with experimental growth data and enzyme kinetic assays, we mapped the sulfur metabolic network in N. gonorrhoeae. We confirm that direct acquisition of cystine (and cysteine) supports growth and is essential for colonization of epithelial cells under aerobic conditions. In the absence of cysteine, N. gonorrhoeae exhibits a limited capacity to utilize alternative sulfur sources, with only glutathione supporting proliferation, albeit inefficiently, via an unidentified mechanism. Growth under these conditions was restored by further supplementation with thiosulfate or methionine, indicating that glutathione utilization is constrained by downstream metabolic limitations.
This limited capacity to utilize alternative sulfur acquisition strategies is notable given that cysteine availability within the host is tightly regulated and maintained at low concentrations. Transcriptomic analyses across cysteine-independent growth conditions revealed the sulfur landscape influences transcriptional responses in a metabolite specific manner. More specifically, in response to sulfur starvation, N. gonorrhoeae does not appear to overcome this limitation through diverse acquisition strategies but instead adjusts cellular processes to compensate for inefficient sulfur acquisition via resource reallocation, where possible.
By defining the sulfur metabolic network and uncovering the transcriptional responses, we conclude that N. gonorrhoeae exhibits metabolic inflexibility when its preferred substrate, cysteine, is unavailable, with proliferation supported by condition-specific adaptive responses rather than broad metabolic versatility. These findings provide valuable insights into how the gonococcus adapts to nutrient scarcity and advances our understanding of the fundamental physiology of this host-adapted pathogen.