Submitted to: Microbiological Research
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 8/31/2015
Publication Date: 1/16/2016
Citation: Gilbert, M.K., Mack, B.M., Wei, Q., Bland, J.M., Bhatnagar, D., Cary, J.W. 2016. RNA sequencing of an nsdC mutant reveals global regulation of secondary metabolic gene clusters in Aspergillus flavus. Microbiological Research. 182:150-161.
Interpretive Summary: The filamentous fungi Aspergillus flavus has the potential to infect and contaminate several crops, including corn, peanuts and cotton. Once infection occurs the fungus releases several toxins that pose a major health threat to plant and animal consumers of the crop. Identifying and characterizing the genes responsible for producing these toxins, and other secondary metabolites, remains a priority. One such gene, nsdC, has been shown to be vital for the production of the most carcinogenic and abundant of the toxins, aflatoxin. It is also has a role in the development of sexual and asexual reproductive structures. Here we conducted RNA-sequencing analysis on an nsdC knockout mutant and analyze its influence on the transcription of secondary metabolites. We show that nsdC maintains its importance for aflatoxin production in the high aflatoxin producing strain Af70. We also identify the primary targets of nsdC to be metabolic enzymes. We demonstrate that nsdC is necessary for the production of a number of other metabolites, including penicillin and aflatrem at the gene level. We show experimentally that aflatrem production is decreased in the knockout mutant. This work provides new working knowledge of what is clearly a globally acting transcription factor and essential regulator in both sexual/asexual development and secondary metabolism.
Technical Abstract: The zinc finger transcription factor nsdC is required for both sexual development and aflatoxin production in the saprophytic fungus Aspergillus flavus. While previous work with an nsdC knockout mutant was conducted in Aspergillus nidulans and A. flavus strain 3357, here we demonstrate perturbations in conidiophore development, sclerotial development and a decrease in aflatoxin production in A. flavus strain Af70, a high producer of aflatoxin. We conducted RNA sequencing analysis of the knockout mutant and demonstrate that many genes under regulation by nsdC are related to secondary metabolic clusters, including aflatoxin, penicillin and aflatrem. Further, HPLC analyses demonstrated a decrease in aflatrem levels. Using SMURF analysis combined with RNA sequencing data we illustrate the discovery of a novel metabolic cluster present in Af70 that is not present in strain 3357 that is also regulated by nsdC. Taken together this data identifies nsdC as a globally acting transcription factor that is an essential regulator of both sexual/asexual reproduction and secondary metabolic clusters.