NIRA, THE PATHWAY-SPECIFIC REGULATORY GENE OF NITRATE ASSIMILATION IN ASPERGILLUS-NIDULANS, ENCODES A PUTATIVE GAL4-TYPE ZINC FINGER PROTEIN AND CONTAINS 4 INTRONS IN HIGHLY CONSERVED REGIONS
NIRA, THE PATHWAY-SPECIFIC REGULATORY GENE OF NITRATE ASSIMILATION IN ASPERGILLUS-NIDULANS, ENCODES A PUTATIVE GAL4-TYPE ZINC FINGER PROTEIN AND CONTAINS 4 INTRONS IN HIGHLY CONSERVED REGIONS
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DOI：
10.1128/mcb.11.11.5746
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发表时间：
1991-11-01
影响因子：
5.3
通讯作者：
LANG, BF
中科院分区：
文献类型：
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作者：
BURGER, G;STRAUSS, J;LANG, BF
已经确定了在构巢曲霉中介导硝酸盐诱导的 nirA 的核苷酸序列。 cDNA和基因组DNA序列的比对表明该基因含有四个内含子并编码892个氨基酸的蛋白质。推导的 NIRA 蛋白显示出转录激活因子的所有特征。氨基末端部分的假定双链 DNA 结合结构域包含六个半胱氨酸残基，这是锌指蛋白 GAL4 家族的特征。还存在氨基末端高酸性区域和两个富含脯氨酸的区域。两个突变的核苷酸序列在通过重叠DNA片段转化并通过聚合酶链式反应扩增后被确定。 nirA87 是一种不可被硝酸盐和亚硝酸盐诱导的突变，在三联体 340 处发生 -1 移码，从而消除了多肽中的 549 个 C 端氨基酸。假设截短的多肽是稳定的，它包含锌指结构域和酸性区域，这似乎不足以用于转录激活。 nirA(d)-106 是一种赋予氮代谢物去抑制硝酸盐和亚硝酸盐还原酶活性的等位基因，包括两个转变，将谷氨酸变为赖氨酸，将缬氨酸变为丙氨酸，位于蛋白质的碱性区域和富含脯氨酸区域之间。对野生型以及组成型 (nirA(c)) 和去阻抑型 (nirA(d)) 突变体的 Northern (RNA) 分析表明，nirA 转录物在这些菌株之间没有变化，在所有情况下都是组成型表达。另一方面，结构基因（niaD 和 niiA）的转录水平确实有所不同，在野生型中高度诱导，但在 nirA(c) 突变体中组成型表达。 nirA(d) 突变体表现出表型去抑制，因为 niaD 和 niiA 转录水平在硝酸盐存在下被过度诱导，但在铵存在下仍然部分受到抑制。
The nucleotide sequence of nirA, mediating nitrate induction in Aspergillus nidulans, has been determined. Alignment of the cDNA and the genomic DNA sequence indicates that the gene contains four introns and encodes a protein of 892 amino acids. The deduced NIRA protein displays all characteristics of a transcriptional activator. A putative double-stranded DNA-binding domain in the amino-terminal part comprises six cysteine residues, characteristic for the GAL4 family of zinc finger proteins. An amino-terminal highly acidic region and two proline-rich regions are also present. The nucleotide sequences of two mutations were determined after they were mapped by transformation with overlapping DNA fragments, amplified by the polymerase chain reaction. nirA87, a mutation conferring noninducibility by nitrate and nitrite, has a -1 frameshift at triplet 340, which eliminates 549 C-terminal amino acids from the polypeptide. Under the assumption that the truncated polypeptide is stable, it comprises the zinc finger domain and the acidic region, which seem not sufficient for transcriptional activation. nirA(d)-106, an allele conferring nitrogen metabolite derepression of nitrate and nitrite reductase activity, includes two transitions, changing a glutamic acid to a lysine and a valine to an alanine, situated between a basic and a proline-rich region of the protein. Northern (RNA) analysis of the wild type and of constitutive (nirA(c)) and derepressed (nirA(d)) mutants show that the nirA transcript does not vary between these strains, being in all cases constitutively expressed. On the other hand, transcript levels of structural genes (niaD and niiA) do vary, being highly inducible in the wild type but constitutively expressed in the nirA(c) mutant. The nirA(d) mutant appears phenotypically derepressed, because the niaD and niiA transcript levels are overinduced in the presence of nitrate but are still partially repressed in the presence of ammonium.