Sexual Development and Meiotic Silencing in Neurospora crassa

Abstract

Meiotic silencing refers to the mechanism of silencing genes or chromosomes without a homologous counterpart (unpaired) during meiotic prophase I. Meiotic silencing has been described in several eukaryotes, including humans. Failure to complete meiotic silencing may be detrimental to the organism. There is a necessity for understanding the regulation of the process. Neurospora is a powerful model system to study gene silencing phenomena. Numerous genes have been determined to be involved in meiotic silencing in Neurospora; however, very little is known about the molecular mechanisms underlying the process. To understand the regulation of meiotic silencing, it is required to combine different approaches such as genetics, proteomics and biochemical analyses.

There is a need for introducing biochemical approaches to the study of meiotic silencing and other processes occurring during sexual development in N. crassa. However, protein extraction from sexual tissue is challenging due to the mechanical difficulties associated with disruption of sexual structure. I standardized a strategy that optimizes protein extraction from sexual tissue. Using this strategy, I studied protein-protein interactions among components of the meiotic silencing machinery and determine the proteome of sexual development.

I identified new protein interactions during meiotic silencing in N. crassa, and established protein-binding partners for the suppressor of meiotic silencing SMS-5. These interacting partners, PAF400 and Pianissimo represent new molecular components involved in the nuclear initial stage of the meiotic silencing mechanism. Interactions between SMS-5, PAF400, and Pianissimo may represent the connection between chromatin remodeling, DNA repair, signaling transduction pathways and meiotic silencing.

I describe the experiments and data analyses used to develop a comprehensive proteomics data set and a functional catalogue for N. crassa sexual development. I used a global proteomics approach and comparative protein functional analysis to investigate the potential molecular differences between two stages of sexual development in filamentous fungi. The data show that secondary metabolites biosynthesis and cellulase activity are required in fruiting body maturation. N. crassa functional catalogue of sexual development proteins will serve as a reference tool for further studies related to sexual development not only in N. crassa, but also in other filamentous ascomycetes.