Molecular cloning and analysis of the regulation of cys-14+, a structural gene of the sulfur regulatory circuit of Neurospora crassa.

Ketter, James S.; Marzluf, George A.

doi:10.1128/mcb.8.4.1504

Cited by 39 publications

(29 citation statements)

References 26 publications

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“…In either case, the effect of the regulatory mutant was to increase ars-l+ message content. It is of interest that cys-14+ (sulfate permease) is affected similarly by cys-3 and sconc with regard to message content (19).…”

Section: Resultsmentioning

confidence: 99%

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Molecular cloning and regulatory analysis of the arylsulfatase structural gene of Neurospora crassa.

Paietta

1989

Mol. Cell. Biol.

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The ars-l+ gene of Neurospora crassa encodes the enzyme arylsulfatase. ars-l+ is in a group of highly regulated sulfur-related structural genes that are expressed under conditions of sulfur limitation and are under coordinate control of the cys-3+ and scon+ regulatory genes. The ars-l+ gene was cloned by chromosome walking from the qa gene cluster, using a lambda library. Cotransformation of an N. crassa ars-1 mutant with the isolated lambda clones and the benomyl resistance gene, followed by assay for arylsulfatase activity, was used to screen for the ars-l+ gene. Further confirmation that the cloned segment mapped to the ars-l+ locus was obtained by restriction-fragment-length polymorphism analysis. Northern (RNA) blot analysis showed that the ars-l + gene was transcribed to give an mRNA of 2.3 kilobases. In wild-type cells, the ars-1 + transcript was abundant under sulfur-derepressing conditions but absent under repressing conditions. Time course analysis showed that the appearance of ars-l+ message in sulfur-derepressed cultures paralleled the appearance of arylsulfatase enzyme activity. In addition, transcription of ars-l+ was detected only under derepressing conditions in a nuclear transcription assay. In a cys-3 regulatory mutant that was unable to synthesize arylsulfatase (or other sulfur-controlled enzymes), there was no ars-l+ transcript under repressing or derepressing conditions. In a temperature-sensitive cys-3 mutant, the ars-l + transcript was present only at the permissive growth temperature and under sulfur derepression. A negative regulatory mutant, sconC, displayed both constitutive expression of arylsulfatase enzyme activity and content of ars-l + message.In the filamentous fungus Neurospora crassa, the synthesis of sets of enzymes in global areas of metabolism such as nitrogen, phosphorus, and sulfur metabolism is highly regulated (24, 26). The sulfur regulatory circuit of N. crassa provides a model system for studying coordinate gene regulation and how a cell regulates its sulfur status. The sulfur control system is composed of a genetically defined set of trans-acting regulatory genes and a set of structural genes that encode enzymes used in the uptake and assimilation of a variety of sulfur compounds (6,26). When cultures of N. crassa are grown under conditions of sulfur limitation (i.e., derepressing conditions), then the set of sulfur-related genes is expressed in a coordinate manner. The structural genes involved encode for arylsulfatase, choline sulfatase, choline sulfate permease, methionine permease, sulfate permeases I and II, and an extracellular protease. None of the known genes in the sulfur regulatory circuit are linked (14,19,23,36).An important part of the sulfur control system involves the positive regulatory gene, cys-3+. In cys-3 mutants there is a pleiotropic loss of the entire set of sulfur-controlled enzymes (25). Paietta et al. (34) cloned the cys-3+ regulatory gene, and subsequent work (12) has shown that the encoded product is a 25.9-kilodalton protein that contains ...

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Section: Resultsmentioning

confidence: 99%

“…The structural genes involved encode for arylsulfatase, choline sulfatase, choline sulfate permease, methionine permease, sulfate permeases I and II, and an extracellular protease. None of the known genes in the sulfur regulatory circuit are linked (14,19,23,36).…”

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Molecular cloning and regulatory analysis of the arylsulfatase structural gene of Neurospora crassa.

Paietta

1989

Mol. Cell. Biol.

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“…Changes in mRNA levels, induced by sulfate starvation, ct,rresponded with changes of the transportation activities determined physiologically [4][5][6][7]. In the filamentous fungus, A eurospora crassa, expression of the cys14 gene encoding sulf~ te permease II was activated by sulfur limitation [8,9]. The p, ~sitive acting regulatory protein, Cys3, which binds to speci iic sites of the 5'-untranslated region of cys14, was required fi,r its expression [9,10].…”

Section: Introductionmentioning

confidence: 99%

Isolation and characterization of a cDNA encoding a sulfate transporter from Arabidopsis thaliana

et al. 1996

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A cDNA encoding a sulfate transporter was isolated from Arabidopsis thaliana. The isolated clone contained an open reading frame encoding a polypeptide of 658 amino acids, exhibiting the highest similarity (62%) with the sequence of the low-affinity sulfate transporter of a tropical legume Stylosanthes hamata. Northern blot analysis indicated the constitutive accumulation of a 2.6 kb length transcript in leaves and roots of seedlings. We also propose that A. thaliana contains three sulfate transporter genes which are expressed as 3.0, 2.7 and 2.6 kb length transcripts, respectively, in an organ-specific manner.

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“…The structural gene for sulfate permease II, cys-14, has been cloned and shown to encode an mRNA of approximately 3 kilobases (kb) whose content is highly regulated by cys-3, by scon, and by the sulfur status of the celis (11). Thus, it appears that both of the regulatory genes as well as sulfur repression act at the level of transcription or at a closely related step such as mRNA processing or stability.…”

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cys-3, the positive-acting sulfur regulatory gene of Neurospora crassa, encodes a protein with a putative leucine zipper DNA-binding element.

Fu¹,

Paietta²,

Mannix³

et al. 1989

Mol. Cell. Biol.

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The sulfur-regulatory circuit of Neurospora crassa consists of a set of unlinked structural genes which encode sulfur-catabolic enzymes and two major regulatory genes which govern their expression. The positive-acting cys-3 regulatory gene is required to turn on the expression of the sulfur-related enzymes, whereas the other regulatory gene, scon, acts in a negative fashion to repress the synthesis of the same set of enzymes. Expression of the cys-3 regulatory gene was found to be controlled by scon and by sulfur availability. The nucleotide sequence of the cys-3 gene was determined and can be translated to yield a protein of molecular weight 25,892 which displays significant homology with the oncogene protein Fos, yeast GCN4 protein, and sea urchin histone Hl. Moreover, the putative cys-3 protein has a well-defined leucine zipper element plus an adjacent charged region which together may make up a DNA-binding site. A cys-3 mutant and a cys-3 temperature-sensitive mutant lead to substitutions of glutamine for basic amino acids within the charged region and thus may alter DNA-binding properties of the cys-3 protein.In the filamentous fungus Neurospora crassa, a high degree of genetic and metabolic regulation governs the expression of sets of enzymes within various global areas of metabolism such as nitrogen, phosphorus, and sulfur catabolism (7, 18). The sulfur control circuit of N. crassa consists of a set of unlinked structural genes which specify enzymes involved in sulfur metabolism. Synthesis of this entire family of sulfur-related enzymes, which includes aryl sulfatase, choline sulfatase, sulfate permease, a high-affinity methionine permease, and an extracellular protease occurs only when cellular levels of sulfur become limited (10,16,19,22). The expression of these sulfur-catabolic enzymes is controlled by two distinct regulatory genes. One of these, designated scon (for sulfur controller), appears to act in a negative fashion; scon mutants are insensitive to sulfur catabolite repression and thus express the sulfur-related enzymes in a constitutive fashion (4). The other sulfurregulatory gene, known as cys-3, acts in a positive manner to activate the expression of the various sulfur-related genes (17,21).Two different sulfate permease species, specified by distinct and unlinked structural genes, are both members of the sulfur circuit (16). The structural gene for sulfate permease II, cys-14, has been cloned and shown to encode an mRNA of approximately 3 kilobases (kb) whose content is highly regulated by cys-3, by scon, and by the sulfur status of the celis (11). Thus, it appears that both of the regulatory genes as well as sulfur repression act at the level of transcription or at a closely related step such as mRNA processing or stability. The cys-3 major sulfur control gene has been postulated to encode a regulatory protein which is needed to turn on the expression of the entire set of sulfur-related activities, presumably by binding at target DNA sequences adjacent to each structural gene (11). Mutants...

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Molecular cloning and analysis of the regulation of cys-14+, a structural gene of the sulfur regulatory circuit of Neurospora crassa.

Cited by 39 publications

References 26 publications

Molecular cloning and regulatory analysis of the arylsulfatase structural gene of Neurospora crassa.

Molecular cloning and regulatory analysis of the arylsulfatase structural gene of Neurospora crassa.

Isolation and characterization of a cDNA encoding a sulfate transporter from Arabidopsis thaliana

cys-3, the positive-acting sulfur regulatory gene of Neurospora crassa, encodes a protein with a putative leucine zipper DNA-binding element.

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