An upstream activating sequence from the Aspergillus nidulans gpdA gene

Punt, Peter J.; Kramer, C.; Kuyvenhoven, A.; Pouwels, Peter H.; Hondel, C.A.M.J.J. van den

doi:10.1016/0378-1119(92)90010-m

Cited by 46 publications

(19 citation statements)

References 29 publications

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“…To verify that the observed defect is due to deletion of the pro45 gene, we ectopically integrated wild-type pro45 gene copies into the ⌬pro45 mutant. Since whole transcriptome shotgun sequencing (RNA-seq) analysis showed that pro45 transcript levels were very low (58), we expressed pro45 from the Aspergillus nidulans gpd promoter (59). Two different translational fusions of PRO45 with a tandem affinity purification (TAP) tag (pro45-TAP) and a GFP tag (EGFP-pro45) were able to complement the sexual developmental defect of the ⌬pro45 mutant ( Fig.…”

Section: Resultsmentioning

confidence: 99%

A Fungal Sarcolemmal Membrane-Associated Protein (SLMAP) Homolog Plays a Fundamental Role in Development and Localizes to the Nuclear Envelope, Endoplasmic Reticulum, and Mitochondria

et al. 2015

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bSarcolemmal membrane-associated protein (SLMAP) is a tail-anchored protein involved in fundamental cellular processes, such as myoblast fusion, cell cycle progression, and chromosomal inheritance. Further, SLMAP misexpression is associated with endothelial dysfunctions in diabetes and cancer. SLMAP is part of the conserved striatin-interacting phosphatase and kinase (STRIPAK) complex required for specific signaling pathways in yeasts, filamentous fungi, insects, and mammals. In filamentous fungi, STRIPAK was initially discovered in Sordaria macrospora, a model system for fungal differentiation. Here, we functionally characterize the STRIPAK subunit PRO45, a homolog of human SLMAP. We show that PRO45 is required for sexual propagation and cell-to-cell fusion and that its forkhead-associated (FHA) domain is essential for these processes. Protein-protein interaction studies revealed that PRO45 binds to STRIPAK subunits PRO11 and SmMOB3, which are also required for sexual propagation. Superresolution structured-illumination microscopy (SIM) further established that PRO45 localizes to the nuclear envelope, endoplasmic reticulum, and mitochondria. SIM also showed that localization to the nuclear envelope requires STRIPAK subunits PRO11 and PRO22, whereas for mitochondria it does not. Taken together, our study provides important insights into fundamental roles of the fungal SLMAP homolog PRO45 and suggests STRIPAK-related and STRIPAK-unrelated functions. Membrane recruitment of protein complexes, cell signaling modules, and enzymes is a critical step for many cellular functions. The family of tail-anchored proteins is recognized for anchoring proteins and vesicles to specific membranes, such as the endoplasmic reticulum (ER) and the outer mitochondrial membrane (1), and tail-anchored proteins are characterized by a C-terminal single transmembrane domain, which is posttranslationally inserted into membranes (2, 3).Sarcolemmal membrane-associated protein (SLMAP) is a tailanchored protein first identified in myocardiac cells (4). In mammals, this protein is known to be involved in myoblast fusion during embryonic development, excitation-contraction coupling in cardiac myocytes, and cell cycle progression (5-8). Furthermore, SLMAP was identified to be a disease gene for Brugada syndrome, a cardiac channelopathy (9). The functional diversity of SLMAP is dependent on alternative splicing, leading to at least four different isoforms of the protein (4, 6, 7, 10). Importantly, gene expression analyses have implicated SLMAP misexpression with endothelial dysfunctions in diabetes, chromosomal aberrations, and cancer (11-14), and currently, SLMAP is the target of lectin-based treatment of drug-resistant cancer cells (15).SLMAP is conserved from yeasts to humans, and characterized fungal SLMAP homologs include Neurospora crassa HAM-4 (hyphal anastomosis 4), Saccharomyces cerevisiae Far9p (factor arrest 9p) and Far10p, as well as Schizosaccharomyces pombe Csc1p (component of SIP complex 1p) (16-18). HAM-4 is essential for vegetativ...

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Section: Resultsmentioning

confidence: 99%

A Fungal Sarcolemmal Membrane-Associated Protein (SLMAP) Homolog Plays a Fundamental Role in Development and Localizes to the Nuclear Envelope, Endoplasmic Reticulum, and Mitochondria

et al. 2015

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“…Integration of wildtype copies of mik1 , mek1 , and mak1 into the respective deletion strains partially complemented the growth defect. This result may be explained by mis-expression of the kinase genes from the constitutive A. nidulans gpd promoter ( mik1 and mek1 ) and the inducible S. macrospora Smxyl promoter ( mak1 ) [38], [39].…”

Section: Resultsmentioning

confidence: 99%

PRO40 Is a Scaffold Protein of the Cell Wall Integrity Pathway, Linking the MAP Kinase Module to the Upstream Activator Protein Kinase C

et al. 2014

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Mitogen-activated protein kinase (MAPK) pathways are crucial signaling instruments in eukaryotes. Most ascomycetes possess three MAPK modules that are involved in key developmental processes like sexual propagation or pathogenesis. However, the regulation of these modules by adapters or scaffolds is largely unknown. Here, we studied the function of the cell wall integrity (CWI) MAPK module in the model fungus Sordaria macrospora. Using a forward genetic approach, we found that sterile mutant pro30 has a mutated mik1 gene that encodes the MAPK kinase kinase (MAPKKK) of the proposed CWI pathway. We generated single deletion mutants lacking MAPKKK MIK1, MAPK kinase (MAPKK) MEK1, or MAPK MAK1 and found them all to be sterile, cell fusion-deficient and highly impaired in vegetative growth and cell wall stress response. By searching for MEK1 interaction partners via tandem affinity purification and mass spectrometry, we identified previously characterized developmental protein PRO40 as a MEK1 interaction partner. Although fungal PRO40 homologs have been implicated in diverse developmental processes, their molecular function is currently unknown. Extensive affinity purification, mass spectrometry, and yeast two-hybrid experiments showed that PRO40 is able to bind MIK1, MEK1, and the upstream activator protein kinase C (PKC1). We further found that the PRO40 N-terminal disordered region and the central region encompassing a WW interaction domain are sufficient to govern interaction with MEK1. Most importantly, time- and stress-dependent phosphorylation studies showed that PRO40 is required for MAK1 activity. The sum of our results implies that PRO40 is a scaffold protein for the CWI pathway, linking the MAPK module to the upstream activator PKC1. Our data provide important insights into the mechanistic role of a protein that has been implicated in sexual and asexual development, cell fusion, symbiosis, and pathogenicity in different fungal systems.

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“…8. Schematic representation of the pTRAN3-1A derivatives with deletions of putative regulatory elements (58,59) indicated by squares (xЈ and xЉ) or carrying insertions of one or two copies of a UAS from a different A. nidulans gene (UAS gpdA [45]). The positions of the NirA binding sites are also indicated.…”

Section: Discussionmentioning

confidence: 99%

The Intergenic Region between the Divergently Transcribed niiA and niaD Genes of Aspergillus nidulans Contains Multiple NirA Binding Sites Which Act Bidirectionally

Punt

Strauss

Smit

et al. 1995

Molecular and Cellular Biology

125

100

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The niaD and niiA genes of Aspergillus nidulans, which code, respectively, for nitrate and nitrite reductases, are divergently transcribed, and their ATGs are separated by 1,200 bp. The genes are under the control of the positively acting NirA transcription factor, which mediates nitrate induction. The DNA binding domain of NirA was expressed as a fusion protein with the glutathione S-transferase of Schistosoma japonicum. Gel shift and footprint experiments have shown that in the intergenic region there are four binding sites for the NirA transcription factor. These sites can be represented by the nonpalindromic consensus 5'CTCCGHGG3'. Making use of a bidirectional expression vector, we have analyzed the role of each of the sites in niaD and niiA expression. The sites were numbered from the niiA side. It appeared that site 1 is necessary for the inducibility of niiA only, while sites 2, 3, and to a lesser extent 4 (which is nearer to and strongly affects niaD) act bidirectionally. The results also suggest that of the 10 binding sites for the AreA protein, which mediates nitrogen metabolite repression, those which are centrally located are physiologically important. The insertion of an unrelated upstream activating sequence into the intergenic region strongly affected the expression of both genes, irrespective of the orientation in which the element was inserted.

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An upstream activating sequence from the Aspergillus nidulans gpdA gene

Cited by 46 publications

References 29 publications

A Fungal Sarcolemmal Membrane-Associated Protein (SLMAP) Homolog Plays a Fundamental Role in Development and Localizes to the Nuclear Envelope, Endoplasmic Reticulum, and Mitochondria

A Fungal Sarcolemmal Membrane-Associated Protein (SLMAP) Homolog Plays a Fundamental Role in Development and Localizes to the Nuclear Envelope, Endoplasmic Reticulum, and Mitochondria

PRO40 Is a Scaffold Protein of the Cell Wall Integrity Pathway, Linking the MAP Kinase Module to the Upstream Activator Protein Kinase C

The Intergenic Region between the Divergently Transcribed niiA and niaD Genes of Aspergillus nidulans Contains Multiple NirA Binding Sites Which Act Bidirectionally

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