Baghela, A. | bioRxiv Science

bioRxiv2021

Deciphering the differential physiological and molecular requirements for conidial anastomosis tube fusion and germ tube formation in Colletotrichum gloeosporioides.

The conidia of a hemibiotrophic fungus Colletotrichum gloeosporioides can conventionally form germ tube (GT) and develop in to a fungal colony, while under certain conditions, they tend to get connected with each other through conidial anastomosis tube (CAT) so as to share the nutrients. CAT fusion is believed to be responsible for generation of genetic variations in few asexual fungi, which appears problematic for effective fungal disease management. The physiological and molecular mechanism underlying the GT versus CAT formation remained unexplored. In the present study, we have deciphered the decision switch responsible for GT formation versus CAT fusion in C. gloeosporioides. GT formation occurred at high frequency in the presence of nutrients, while CAT fusion was found to be higher in absence of nutrients. Younger conidia were found to form GT efficiently, whilst older conidia preferentially formed CAT. Whole transcriptome analysis of GT and CAT fusion revealed differential molecular requirements for these two processes. We identified 11050 and 9786 differentially expressed genes (DEGs) in GT and CAT, respectively. A total 1567 effector candidates were identified, of them 103 and 101 were uniquely secreted during GT and CAT formation respectively. Genes coding for cell wall degrading enzymes, germination, hyphal growth, host-fungus interaction and virulence were up-regulated during GT formation. Whilst, genes involved in stress response, cell wall remodelling, membrane transport, cytoskeleton, cell cycle, and cell rescue were highly up-regulated during CAT fusion. To conclude, the GT and CAT fusion were found to be mutually exclusive processes, requiring differential physiological conditions and sets of DEGs in C. gloeosporioides. This will help to understand the basic CAT biology in the genus Colletotrichum.

microbiology↗

bioRxiv2020

The stringent stress response controls proteases and global regulators under optimal growth conditions in Pseudomonas aeruginosa

The bacterial stringent stress response, mediated by the signaling molecule guanosine tetra-phosphate, ppGpp, has recently gained attention as being important during normal cellular growth and as potential new therapeutic target, which warrants detailed mechanistic understanding. Here, we used intracellular protein tracking in Pseudomonas aeruginosa PAO1, which indicated that RelA was bound to the ribosome, while SpoT localized at the cell poles. RNA-Seq was used to investigate the transcriptome of a ppGpp-deficient strain under non-stressful nutrient-rich broth conditions where the mutant grew at the same rate as the parent strain. In exponential growth phase, the lack of ppGpp led to >1,600 transcriptional changes (fold-change cut-off {+/-}1.5), providing further novel insights into the normal physiological role of ppGpp. The stringent response was linked to gene expression of various proteases and secretion systems including aprA, PA0277, impA, and clpP2. The previously observed reduction in cytotoxicity towards red blood cells, in a stringent response mutant, appeared to be due to aprA. Investigation of an aprA mutant in a murine skin infection model, showed increased survival rates of the aprA mutant consistent with previous observations that stringent-response mutants have reduced virulence. In addition, the overexpression of relA, but not induction of ppGpp with serine hydroxamate, dysregulated global transcriptional regulators as well as >30% of the regulatory networks controlled by AlgR, OxyR, LasR, and AmrZ. Together these data expand our knowledge about ppGpp and its regulatory network and role in environmental adaptation. It also confirms its important role throughout the normal growth cycle of bacteria. Significance StatementMicroorganisms need to adapt rapidly to survive harsh environmental changes. Here, we showed the broad influence of the highly studied bacterial stringent stress response under non-stressful conditions that indicate its general physiological importance and might reflect the readiness of bacteria to respond to and activate acute stress responses. Using RNA-Seq to investigate the transcriptional network of Pseudomonas aeruginosa cells revealed that >30% of all genes changed expression in a stringent-response mutant under optimal growth conditions. This included genes regulated by global transcriptional regulators and novel downstream effectors. Our results help to understand the importance of this stress regulator in bacterial lifestyle under relatively unstressed conditions. As such it draws attention to the consequences of targeting this ubiquitous bacterial signaling molecule.

microbiology↗