A moonlighting function of a chitin polysaccharide monooxygenase, CWR-1, in Neurospora crassa allorecognition

Abstract

Organisms require the ability to differentiate themselves from organisms of different or even the same species. Allorecognition processes in filamentous fungi are essential to ensure identity of an interconnected syncytial colony to protect it from exploitation and disease. Neurospora crassa has three cell fusion checkpoints controlling formation of an interconnected mycelial network. The locus that controls the second checkpoint, which allows for cell wall dissolution and subsequent fusion between cells/hyphae, cwr (cell wall remodeling), encodes two linked genes, cwr-1 and cwr-2. Previously, it was shown that cwr-1 and cwr-2 show severe linkage disequilibrium with six different haplogroups present in N. crassa populations. Isolates from an identical cwr haplogroup show robust fusion, while somatic cell fusion between isolates of different haplogroups is significantly blocked in cell wall dissolution. The cwr-1 gene encodes a putative polysaccharide monooxygenase (PMO). Herein we confirm that CWR-1 is a C1-oxidizing chitin PMO. We show that the catalytic (PMO) domain of CWR-1 was sufficient for checkpoint function and cell fusion blockage; however, through analysis of active-site, histidine-brace mutants, the catalytic activity of CWR-1 was ruled out as a major factor for allorecognition. Swapping a portion of the PMO domain (V86 to T130) did not switch cwr haplogroup specificity, but rather cells containing this chimera exhibited a novel haplogroup specificity. Allorecognition to mediate cell fusion blockage is likely occurring through a protein-protein interaction between CWR-1 with CWR-2. These data highlight a moonlighting role in allorecognition of the CWR-1 PMO domain.

A fungal lytic polysaccharide monooxygenase is required for cell wall integrity, thermotolerance, and virulence of the fungal human pathogen Cryptococcus neoformans

“…The CWR-1 and CWR-2 proteins were shown to be required for recognizing strains of different haplotypes that were compatible for filament fusion. Interestingly, and in contrast to our findings in Cn Cel1, this activity was independent of its catalytic function as mutation of the copper-binding residues did not affect this aspect of protein function [ 62 ].…”

A fungal lytic polysaccharide monooxygenase is required for cell wall integrity, thermotolerance, and virulence of the fungal human pathogenCryptococcus neoformans

“…Phenotypic analysis of the cel1 mutant suggests that this protein is involved in cell wall strains of different haplotypes that were compatible for filament fusion. Interestingly, and in contrast to our findings in CnCel1, this activity was independent of its catalytic function as mutation of the copper-binding residues did not affect this aspect of protein function [62].…”

AA16 Oxidoreductases Boost Cellulose-Active AA9 Lytic Polysaccharide Monooxygenases from Myceliophthora thermophila

“…It might be because Mt AA16A and An AA16A form weak transient interactions with the Mt LPMO9s, which is challenging to study as reviewed by Qin et al 62 However, of note here is that in a recent study, protein–protein interaction between a cell wall remodeling (CWR) protein CWR-1 AA11 LPMO and a CWR-2 membrane protein was implicated to be important for allorecognition of N. crassa . 63 Figure S19 shows a model of a hypothetical protein–protein interaction between Mt LPMO9B and Mt AA16A. It should be emphasized that this model, though having a high score, only represents an illustrative model and other factors such as glycosylation location, CBM, and linker were not taken into account.…”