Large-Scale Transcriptional Response to Hypoxia in Aspergillus fumigatus Observed Using RNAseq Identifies a Novel Hypoxia Regulated ncRNA

Abstract

We utilized RNAseq analysis of the Aspergillus fumigatus response to early hypoxic condition exposure. The results show that more than 89% of the A. fumigatus genome is expressed under normoxic and hypoxic conditions. Replicate samples were highly reproducible; however, comparisons between normoxia and hypoxia revealed that greater than 23% and 35% of genes were differentially expressed after 30 and 120 minutes of hypoxia exposure, respectively. Consistent with our previous report detailing transcriptomic and proteomic responses at later time points, the results here show major repression of ribosomal function and induction of ergosterol biosynthesis, as well as activation of alternate respiratory mechanisms at the later time point. RNAseq data was used to define thirty-two hypoxia-specific genes, which are not expressed under normoxic conditions. Transcripts of a C6 transcription factor and a histidine kinase response regulator were found only in hypoxia. In addition, several genes involved in the phosphoenyl pyruvate (PEP) and D-glyceraldehyde-3-phosphate (G3P) metabolism were only expressed in hypoxia. Interestingly, a 216 bp ncRNA Afu-182 in the 3’ region of insA (AFUB_064770), was significantly repressed under hypoxia with a 40-fold reduction in expression. A detailed analysis of Afu-182 showed similarity with several genes in the genome, many of which were also repressed in hypoxia. The results from this study show that hypoxia induces very early and widely drastic genome-wide responses in A. fumigatus that include expression of protein-coding and ncRNA genes. The role of these ncRNA genes in regulating the fungal hypoxia response is an exciting future research direction.

Two C4-sterol methyl oxidases (Erg25) catalyse ergosterol intermediate demethylation and impact environmental stress adaptation in Aspergillus fumigatus

“…Experimental evidence suggested that AFUB_099650 mRNA increased dramatically when exposed to hypoxia when using nitrate as its sole nitrogen source (Losada et al, 2014;Schinko et al, 2010;Zhou et al, 2009). pflavA-erg25A : : DsrbA was verified to have a single insertion by PCR and Southern blot analysis (Fig.…”

Insights into the cellular responses to hypoxia in filamentous fungi

“…At present, genome-wide transcriptional profiling data of the hypoxic stress response are available from several filamentous ascomycetes: A. nidulans, A. oryzae, A. fumigatus, and Trichoderma reesei (Barker et al 2012;Bonaccorsi et al 2006;Hillmann et al 2014;Kroll et al 2014;Losada et al 2014;Masuo et al 2010;Terabayashi et al 2012). A somewhat smaller number of studies have been published on proteomic changes of A. nidulans and A. fumigatus during adaptation to low oxygen partial pressures (Barker et al 2012;Shimizu et al 2009;Vödisch et al 2011).…”

“…Later this proteomic study was complemented by a microarray experiment, which revealed in addition an altered cell wall, polysaccharide metabolism (many induced glucanases) and an increased metal ion transport (mitochondrial transporter, copper transport) upon hypoxia . Three further transcriptome/proteome surveys studied the batch growth of A. fumigatus at low O 2 levels in an oxygen-controlled fermenter (Barker et al 2012;Hillmann et al 2014;Losada et al 2014). The exposure time to hypoxia varied between 1, 2, and 24 h. Altogether, the number and type of genes whose expression changed significantly in abundance during hypoxic growth differed appreciably between the three studies.…”

Advances Against Aspergillosis: Biology, Host response, Diagnosis and Treatment

“…During infection, A. fumigatus must also adapt to the oxygen-limited environment of host tissues [7]. Losada et al [8] report the results of a study to determine the genome-wide expression profile of A. fumigatus exposed to a hypoxic environment. Using RNA-seq technology, the authors expand previous microarray data on the hypoxia transcriptome and provide intriguing evidence for the existence of a novel hypoxia-regulated non-coding RNA [8].…”

“…Losada et al [8] report the results of a study to determine the genome-wide expression profile of A. fumigatus exposed to a hypoxic environment. Using RNA-seq technology, the authors expand previous microarray data on the hypoxia transcriptome and provide intriguing evidence for the existence of a novel hypoxia-regulated non-coding RNA [8]. Another central mediator of eukaryotic stress responses is the calcium/calmodulin-activated protein phosphatase calcineurin, which controls a pathway that regulates drug tolerance, morphogenesis, and virulence in a variety of pathogenic fungi, including A. fumigatus [9].…”