Functional characterization of a Penicillium chrysogenum mutanase gene induced upon co-cultivation with Bacillus subtilis

Abstract

BackgroundMicrobial gene expression is strongly influenced by environmental growth conditions. Comparison of gene expression under different conditions is frequently used for functional analysis and to unravel regulatory networks, however, gene expression responses to co-cultivation with other microorganisms, a common occurrence in nature, is rarely studied under laboratory conditions. To explore cellular responses of the antibiotic-producing fungus Penicillium chrysogenum to prokaryotes, the present study investigates its transcriptional responses during co-cultivation with Bacillus subtilis.ResultsSteady-state glucose-limited chemostats of P. chrysogenum grown under penillicin-non-producing conditions were inoculated with B. subtilis. Physiological and transcriptional responses of P. chrysogenum in the resulting mixed culture were monitored over 72 h. Under these conditions, B. subtilis outcompeted P. chrysogenum, as reflected by a three-fold increase of the B. subtilis population size and a two-fold reduction of the P. chrysogenum biomass concentration. Genes involved in the penicillin pathway and in synthesis of the penicillin precursors and side-chain were unresponsive to the presence of B. subtilis. Moreover, Penicillium polyketide synthase and nonribosomal peptide synthase genes were either not expressed or down-regulated. Among the highly responsive genes, two putative α-1,3 endoglucanase (mutanase) genes viz Pc12g07500 and Pc12g13330 were upregulated by more than 15-fold and 8-fold, respectively. Measurement of enzyme activity in the supernatant of mixed culture confirmed that the co-cultivation with B. subtilis induced mutanase production. Mutanase activity was neither observed in pure cultures of P. chrysogenum or B. subtilis, nor during exposure of P. chrysogenum to B. subtilis culture supernatants or heat-inactivated B. subtilis cells. However, mutanase production was observed in cultures of P. chrysogenum exposed to filter-sterilized supernatants of mixed cultures of P. chrysogenum and B. subtilis. Heterologous expression of Pc12g07500 and Pc12g13330 genes in Saccharomyces cerevisiae confirmed that Pc12g07500 encoded an active α-1,3 endoglucanase.ConclusionTime-course transcriptional profiling of P. chrysogenum revealed differentially expressed genes during co-cultivation with B. subtilis. Penicillin production was not induced under these conditions. However, induction of a newly characterized P. chrysogenum gene encoding α-1,3 endoglucanase may enhance the efficacy of fungal antibiotics by degrading bacterial exopolysaccharides.

Diversity of Filamentous Fungi Isolated From Some Amylase and Alcohol-Producing Starters of India

“…The probable entry of P. chrysogenum during traditional preparation may be from damp and moist rooms where preparation for such starters is usually done, since P. chrysogenum is also found in damp buildings (Andersen et al, 2011). Due to the ability of P. chrysogenum to produce antibiotics, mostly penicillin (Bajaj et al, 2014), its presence in starters may have an antagonist property in the end product. P. citrinum was recovered in samples of humao, hamei and khekhrii, probably from indoor environments (Samson et al, 2004).…”

The low affinity glucose transporter HxtB is also involved in glucose signalling and metabolism in Aspergillus nidulans

“…Furthermore, A. nidulans pellet formation was prevented allowing for homogenous, macroscopic morphology and glucose uptake throughout the culture. Continuous chemostat cultivation has been carried out for several filamentous fungi such as A. niger 34 , T. reesei 47 , Penicillium chrysogenum 48 and N. crassa 49 but until now has not been published for A. nidulans . This study shows that continuous batch and chemostat cultivation can also be applied to A. nidulans , which is considered an important ‘reference’ organism for studying various cellular processes in filamentous fungi.…”

Biomass from phytopathogens and culture conditions improve Penicillium chrysogenum antimicrobial activity and antifungal compounds production

“…Several authors have reported different strains of the Penicillium fungus as controlling the growth of phytopathogenic bacteria and fungi (Anitha & Rabeeth, 2009;Bajaj et al, 2014;Bolívar-Anillo et al, 2020;Da Silva Sousa et al, 2008;Manigundan et al, 2020;Sabuquillo et al, 2006). Some Penicillium strains have been shown to control economically important phytopathogenic fungi such as R. solanacearum (Wattana-Amorn et al, 2016), P. oryzae (Law et al, 2017), and C. dematium (Song et al 2020).…”