Molecular comparison of the negative-acting nitrogen control gene,nmr, inNeurospora crassa and otherNeurospora and fungal species

Abstract

In Neurospora crassa, the expression of unlinked structural genes which encode nitrogen catabolic enzymes is subject to genetic and metabolic regulation. The negative-acting nmr regulatory gene appears to play a role in nitrogen catabolite repression. Using the N. crassa nmr gene as a probe, homologous sequences were identified in a variety of other filamentous fungi. The polymerase chain reaction was used to isolate the nmr-like gene from the exotic Mauriceville strain of N. crassa and from the two related species, N. intermedia and N. sitophila. Sequence comparisons were carried out with a 1.7-kb DNA segment which includes the entire coding region of nmr plus 5' and 3' noncoding sequences. The size of the nmr coding region was identical in all three Neurospora species. Approximately 30 nucleotide base substitutions were found in the coding region of the nmr gene of each of the sister species when compared to the standard N. crassa sequence. However, most of the base changes occurred in third codon positions and were silent. The NMR proteins of N. sitophila and of N. intermedia display only three and four amino acid substitutions, respectively, from the N. crassa protein. Two regions of high variability, which include deletions and insertions of bases, were found in the 5' and 3' noncoding regions of the gene.

Nitrogen metabolite signalling involves the C-terminus and the GATA domain of the Aspergillus transcription factor AREA and the 3′ untranslated region of its mRNA.

“…The role of GATA domains in DNA binding has been clearly demonstrated (e.g. A good candidate would be an Aspergillus homologue (Young and Marzluf, 1991; S.Polley and M.X.Caddick, unpublished data) of NMR1, a putative negative regulatory protein identified in N.crassa (Young et al, 1990 and references therein) which interacts directly with both the C-terminus and the DNA-binding domain of the AREA homologue, NIT2 (Xiao et al, 1995). These include nuclear localization (Yang et al, 1994), transcriptional activation (Martin and Orkin, 1990;Yang and Evans, 1992) and protein-protein interactions (Peters and Caddick, 1994) including the formation of homodimers and the binding of heterologous proteins (Crossley et al, 1995;Merika and Orkin, 1995;Yang and Evans, 1995;Xiao et al, 1995).…”

Neurospora from Natural Populations: A Global Study

“…New examples are cdr (Levine and Marzluf, 1989), mcb and mcm (Maheshwari, 1991), and sen (Navaraj et al, 2000). Other studies involved new alleles at already established loci, for example, a novel nmr allele (Young and Marzluf, 1991). Naturally occurring duplicated and nonduplicated versions of a 5S RNA gene were used in a study relating DNA methylation to duplication (Selker and Stevens, 1987).…”

The Negative Transcriptional Regulator NmrA Discriminates between Oxidized and Reduced Dinucleotides

“…In vivo post-translational modulation of NIT2 activity has been implicated, as in vitro the nmr1-encoded N-terminally deleted forms of NMR1 protein bind directly to the zinc finger region and the extreme C-terminal 30 amino acids of NIT2 (15)(16)(17). The interactions were demonstrated in a qualitative manner by using the yeast two-hybrid system, as well as steady state techniques that included electrophoretic mobility shift assay and column binding using His 6 -and glutathione S-transferasetagged fragments of the NIT2 protein as a bait.…”