TRANSCRIPTION OF THE NEUROSPORA CRASSA HSP70-1 AND HSP70-2 GENES IS DEPENDENT ON THE PACC⁺ BACKGROUND IN AN ACIDIC MILIEU

Fabio M Squina (CTBE); Juliana Leal (FMRP-USP); Vivian Tf Cipriano (FMRP-USP); Nilce M Martinez-rossi (FMRP-USP); Antonio Rossi (FMRP-USP)

Heat shock proteins belong to a conserved superfamily of molecular chaperones found in prokaryotes and eukaryotes. These proteins are linked to a myriad of physiological functions. In this study, we show that the N. crassa hsp70-1 (NCU09602.3) and hsp70-2 (NCU08693.3) genes are preferentially expressed in an acidic milieu after 15 h of cell growth in sufficient phosphate at 30°C. No significant accumulation of these transcripts was detected at alkaline pH values. Both genes accumulated to a high level in mycelia that were incubated for 1 h at 45°C, regardless of the phosphate concentration and extracellular pH changes. Transcription of the hsp70-1 and hsp70-2 genes was dependent on the pacC⁺ background in mycelia cultured under optimal growth conditions or at 45°C. The pacC gene encodes a Zn-finger transcription factor that is involved in the regulation of gene expression by pH. PacC binds to 5'-GCCARG-3' sequences up-stream of pH-conditioned genes and either activates or represses transcription. The initial guanine residue in this consensus sequence is critical for PacC binding, whereas the Rim101p binding sites is 5'-NCCAAG-3' followed preferentially by A or C in the adjacent 3' positions. The presence of this pentanucleotide followed by A or C was identified in the sequences up-stream of the N. crassa hsp70-1, hsp70-2, hsp70-3, hsf2 (NCU08480) and hsf3 (NCU02413) genes, whereas this binding consensus is absent in the sequence up-stream of the N. crassa hsf1 (NCU08512) gene. Thus, hsf1 gene might not be under the direct control of PACC. Heat shock induction of the hsp70-1 and hsp70-2 genes in mycelia incubated in low-phosphate medium was almost not altered in the nuc-1⁻ background under both acidic and alkaline pH conditions. The NUC-1 transcriptional regulator is involved in the derepression of nucleases, phosphatases, and transporters that are necessary for fulfilling the cell's phosphate requirements. Transcription of the hsp70-3 (NCU01499.3) gene followed a different pattern of induction—the gene was depressed under insufficient phosphate conditions but was apparently unaffected by alkalinization of the culture medium. Moreover, this gene was not induced by heat shock. These results reveal novel aspects of the heat-sensing network of N. crassa.