Structure of Saccharomyces cerevisiae mating hormone a-factor. Identification of S-farnesyl cysteine as a structural component.

Anderegg, Robert J.; Betz, Richard; Carr, Steven A.; Crabb, John W.; Duntze, W.

doi:10.1016/s0021-9258(19)81351-0

Cited by 355 publications

(53 citation statements)

References 11 publications

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“…(The [ 3 H]proline signal for M was too low to count.) These results indicate that intracellular M has the same NH 2 terminus (Y22) as that previously established for extracellular M by mass spectrometry (Anderegg et al, 1988); thus, intracellular M has undergone complete removal of the NH 2 -terminal extension.…”

Section: P1 Has An Intact Nh 2 -Terminal Extension P2 Has a Partially Processed Nh 2 -Terminal Extension And M Is Fully Processedsupporting

confidence: 77%

Biogenesis of the Saccharomyces cerevisiae Mating Pheromone a-Factor

Chen

Sapperstein

Choi

et al. 1997

The Journal of Cell Biology

127

177

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The Saccharomyces cerevisiae mating pheromone a-factor is a prenylated and carboxyl methylated extracellular peptide signaling molecule. Biogenesis of the a-factor precursor proceeds via a distinctive multistep pathway that involves COOH-terminal modification, NH2-terminal proteolysis, and a nonclassical export mechanism. In this study, we examine the formation and fate of a-factor biosynthetic intermediates to more precisely define the events that occur during a-factor biogenesis. We have identified four distinct a-factor biosynthetic intermediates (P0, P1, P2, and M) by metabolic labeling, immunoprecipitation, and SDSPAGE. We determined the biochemical composition of each by defining their NH2-terminal amino acid and COOH-terminal modification status. Unexpectedly, we discovered that not one, but two NH2-terminal cleavage steps occur during the biogenesis of a-factor. In addition, we have shown that COOH-terminal prenylation is required for the NH2-terminal processing of a-factor and that all the prenylated a-factor intermediates (P1, P2, and M) are membrane bound, suggesting that many steps of a-factor biogenesis occur in association with membranes. We also observed that although the biogenesis of a-factor is a rapid process, it is inherently inefficient, perhaps reflecting the potential for regulation. Previous studies have identified gene products that participate in the COOH-terminal modification (Ram1p, Ram2p, Ste14p), NH2-terminal processing (Ste24p, Axl1p), and export (Ste6p) of a-factor. The intermediates defined in the present study are discussed in the context of these biogenesis components to formulate an overall model for the pathway of a-factor biogenesis.

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Section: P1 Has An Intact Nh 2 -Terminal Extension P2 Has a Partially Processed Nh 2 -Terminal Extension And M Is Fully Processedsupporting

confidence: 77%

Biogenesis of the Saccharomyces cerevisiae Mating Pheromone a-Factor

Chen

Sapperstein

Choi

et al. 1997

The Journal of Cell Biology

127

177

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“…Lack of the signal peptide and a blocked N-terminus were found to be characteristic for a number of secreted proteins with unknown secretory mechanism, such as the FGFs, PD-ECGF, interleukin 1, parathymosin, prothymosin, ␤ 4-thymosin, and different transglutaminases such as the blood coagulating factor XIII ␣ (Muesch et al 1990). Alternative known pathways, e.g., the ATP-dependent ABC transporters (Higgins 1992) or ␣ -factor-mediated export systems described for yeast cells could be excluded (Anderegg et al 1988). In some of these cases apocrine secretion, which has previously been described in detail for secretory transglutaminase of rat coagulating gland, can be assumed as a potential export pathway (Seitz et al 1990;Steinhoff et al 1994).…”

Section: Discussionmentioning

confidence: 99%

Cytoplasmic Carbonic Anhydrase II of Rat Coagulating Gland Is Secreted via the Apocrine Export Mode

Wilhelm

Keppler

Hoffbauer

et al. 1998

J Histochem Cytochem.

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Two different pathways for protein secretion are described for epithelial cells of rat coagulating gland and dorsal prostate: the classical merocrine and the alternative apocrine release mode. Apocrine-secreted proteins are synthesized on cytoplasmic polyribosomes and are subsequently exported in protrusions on the apical cell surface (aposomes). In this article we report the identification and purification to homogeneity of a 29-kD protein from the secretion of rat coagulating gland. N-terminal amino acid sequence analyses revealed 100% identity to rat brain carbonic anhydrase II (CAH II). In addition, the 29-kD protein showed CAH enzyme activity. On Western blot analysis, a polyclonal anti-CAH II antibody raised in rabbit reacted specifically with the rat and human but not bovine CAH II isoforms. Immunohistochemical studies on rat coagulating gland showed strong labeling for CAH II protein in aposomes. Immunoelectron microscopy confined CAH II protein to the cytoplasm and aposomes, whereas no staining was visible in the compartments of the classical merocrine route, the endoplasmic reticulum and Golgi apparatus. The resident cytoplasmic protein lactate dehydrogenase, however, was not found in the secretion. Taken together, the morphological and biochemical data clearly indicate that cytoplasmic CAH II from rat coagulating gland is specifically selected and then secreted via the apocrine pathway.

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“…The mating pheromone a -factor is encoded by two similar and functionally redundant genes, MFA1 and MFA2 (Brake et al, 1985;Michaelis and Herskowitz, 1988). The a -factor precursors encoded by these genes (36 and 38 amino acids long, respectively) are ultimately processed to the mature, bioactive form of a -factor, which is 12 amino acids long and contains a farnesylated and carboxylmethylated COOH-terminal cysteine residue (Anderegg et al, 1988). A schematic view of the a -factor precursor encoded by MFA1 is shown at the top of Fig.…”

mentioning

confidence: 99%

A Novel Membrane-associated Metalloprotease, Ste24p, Is Required for the First Step of NH2-terminal Processing of the Yeast a-Factor Precursor

Fujimura-Kamada

Nouvet

Michaelis

1997

The Journal of Cell Biology

147

204

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Many secreted bioactive signaling molecules, including the yeast mating pheromones a-factor and α-factor, are initially synthesized as precursors requiring multiple intracellular processing enzymes to generate their mature forms. To identify new gene products involved in the biogenesis of a-factor in Saccharomyces cerevisiae, we carried out a screen for MAT a-specific, mating-defective mutants. We have identified a new mutant, ste24, in addition to previously known sterile mutants. During its biogenesis in a wild-type strain, the a-factor precursor undergoes a series of COOH-terminal CAAX modifications, two sequential NH2-terminal cleavage events, and export from the cell. Identification of the a-factor biosynthetic intermediate that accumulates in the ste24 mutant revealed that STE24 is required for the first NH2-terminal proteolytic processing event within the a-factor precursor, which takes place after COOH-terminal CAAX modification is complete. The STE24 gene product contains multiple predicted membrane spans, a zinc metalloprotease motif (HEXXH), and a COOH-terminal ER retrieval signal (KKXX). The HEXXH protease motif is critical for STE24 activity, since STE24 fails to function when conserved residues within this motif are mutated. The identification of Ste24p homologues in a diverse group of organisms, including Escherichia coli, Schizosaccharomyces pombe, Haemophilus influenzae, and Homo sapiens, indicates that Ste24p has been highly conserved throughout evolution. Ste24p and the proteins related to it define a new subfamily of proteins that are likely to function as intracellular, membrane-associated zinc metalloproteases.

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Structure of Saccharomyces cerevisiae mating hormone a-factor. Identification of S-farnesyl cysteine as a structural component.

Cited by 355 publications

References 11 publications

Biogenesis of the Saccharomyces cerevisiae Mating Pheromone a-Factor

Biogenesis of the Saccharomyces cerevisiae Mating Pheromone a-Factor

Cytoplasmic Carbonic Anhydrase II of Rat Coagulating Gland Is Secreted via the Apocrine Export Mode

A Novel Membrane-associated Metalloprotease, Ste24p, Is Required for the First Step of NH2-terminal Processing of the Yeast a-Factor Precursor

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