The extremely conserved amino terminus of RAD6 ubiquitin-conjugating enzyme is essential for amino-ena rule-dependent protein degradation

John F. Watkins, Patrick Sung, Satya Prakash, Louise Prakash

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71 Citations (Scopus)

Abstract

The RAD6 gene of Saccharomyces cerevisiae encodes a ubiquitin-conjugating enzyme that is requited for DNA repair, damage-induced mutagenesis, and sporulation. In addition, RAD6 mediates the multiubiquitination and degradation of amino-end rule protein substrates. The structure and function of RAD6 have been remarkably conserved during eukaryotic evolution. Here, we examine the role of the extremely conserved amino terminus, which has remained almost invariant among RAD6 homologs from yeast to human. We show that RAD6 is concentrated in the nucleus and that the amino-terminal deletion mutation, rad6Δ1-9, does not alter the location of the protein. The amino-terminal domain, however, is essential for the multiubiquitination and degradation of amino-end rule substrates. In the rad6Δ1-99 mutant, β-galactosidase proteins bearing destabilizing amino-terminal residues become long lived, and purified rad6Δ1-99, protein is ineffective in ubiquitin-protein ligase (E3)-dependent protein degradation in the proteolytic system derived from rabbit reticulocytes. The amino terminus is required for physical interaction of RAD6 with the yeast UBR1-encoded E3 enzyme, as the rad6Δ1-99 protein is defective in this respect. The rad6Δ1-99 mutant is defective in sporulation, shows reduced efficiency of DNA repair, but is proficient in UV mutagenesis. E3-dependent protein degradation by RAD6 could be essential for sporulation and could affect the efficiency of DNA repair.

Original languageEnglish (US)
Pages (from-to)250-261
Number of pages12
JournalGenes and Development
Volume7
Issue number2
StatePublished - 1993
Externally publishedYes

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Ubiquitin-Conjugating Enzymes
Proteolysis
DNA Repair
Mutagenesis
Proteins
Yeasts
Galactosidases
Ubiquitin-Protein Ligases
Sequence Deletion
Reticulocytes
Mutant Proteins
DNA Damage
Saccharomyces cerevisiae
Rabbits
Enzymes
Genes

Keywords

  • Amino-end rule protein
  • RAD6 gene
  • Saccharomyces cerevisiae
  • Ubiquitin-conjugating enzyme

ASJC Scopus subject areas

  • Developmental Biology
  • Genetics

Cite this

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abstract = "The RAD6 gene of Saccharomyces cerevisiae encodes a ubiquitin-conjugating enzyme that is requited for DNA repair, damage-induced mutagenesis, and sporulation. In addition, RAD6 mediates the multiubiquitination and degradation of amino-end rule protein substrates. The structure and function of RAD6 have been remarkably conserved during eukaryotic evolution. Here, we examine the role of the extremely conserved amino terminus, which has remained almost invariant among RAD6 homologs from yeast to human. We show that RAD6 is concentrated in the nucleus and that the amino-terminal deletion mutation, rad6Δ1-9, does not alter the location of the protein. The amino-terminal domain, however, is essential for the multiubiquitination and degradation of amino-end rule substrates. In the rad6Δ1-99 mutant, β-galactosidase proteins bearing destabilizing amino-terminal residues become long lived, and purified rad6Δ1-99, protein is ineffective in ubiquitin-protein ligase (E3)-dependent protein degradation in the proteolytic system derived from rabbit reticulocytes. The amino terminus is required for physical interaction of RAD6 with the yeast UBR1-encoded E3 enzyme, as the rad6Δ1-99 protein is defective in this respect. The rad6Δ1-99 mutant is defective in sporulation, shows reduced efficiency of DNA repair, but is proficient in UV mutagenesis. E3-dependent protein degradation by RAD6 could be essential for sporulation and could affect the efficiency of DNA repair.",
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T1 - The extremely conserved amino terminus of RAD6 ubiquitin-conjugating enzyme is essential for amino-ena rule-dependent protein degradation

AU - Watkins, John F.

AU - Sung, Patrick

AU - Prakash, Satya

AU - Prakash, Louise

PY - 1993

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N2 - The RAD6 gene of Saccharomyces cerevisiae encodes a ubiquitin-conjugating enzyme that is requited for DNA repair, damage-induced mutagenesis, and sporulation. In addition, RAD6 mediates the multiubiquitination and degradation of amino-end rule protein substrates. The structure and function of RAD6 have been remarkably conserved during eukaryotic evolution. Here, we examine the role of the extremely conserved amino terminus, which has remained almost invariant among RAD6 homologs from yeast to human. We show that RAD6 is concentrated in the nucleus and that the amino-terminal deletion mutation, rad6Δ1-9, does not alter the location of the protein. The amino-terminal domain, however, is essential for the multiubiquitination and degradation of amino-end rule substrates. In the rad6Δ1-99 mutant, β-galactosidase proteins bearing destabilizing amino-terminal residues become long lived, and purified rad6Δ1-99, protein is ineffective in ubiquitin-protein ligase (E3)-dependent protein degradation in the proteolytic system derived from rabbit reticulocytes. The amino terminus is required for physical interaction of RAD6 with the yeast UBR1-encoded E3 enzyme, as the rad6Δ1-99 protein is defective in this respect. The rad6Δ1-99 mutant is defective in sporulation, shows reduced efficiency of DNA repair, but is proficient in UV mutagenesis. E3-dependent protein degradation by RAD6 could be essential for sporulation and could affect the efficiency of DNA repair.

AB - The RAD6 gene of Saccharomyces cerevisiae encodes a ubiquitin-conjugating enzyme that is requited for DNA repair, damage-induced mutagenesis, and sporulation. In addition, RAD6 mediates the multiubiquitination and degradation of amino-end rule protein substrates. The structure and function of RAD6 have been remarkably conserved during eukaryotic evolution. Here, we examine the role of the extremely conserved amino terminus, which has remained almost invariant among RAD6 homologs from yeast to human. We show that RAD6 is concentrated in the nucleus and that the amino-terminal deletion mutation, rad6Δ1-9, does not alter the location of the protein. The amino-terminal domain, however, is essential for the multiubiquitination and degradation of amino-end rule substrates. In the rad6Δ1-99 mutant, β-galactosidase proteins bearing destabilizing amino-terminal residues become long lived, and purified rad6Δ1-99, protein is ineffective in ubiquitin-protein ligase (E3)-dependent protein degradation in the proteolytic system derived from rabbit reticulocytes. The amino terminus is required for physical interaction of RAD6 with the yeast UBR1-encoded E3 enzyme, as the rad6Δ1-99 protein is defective in this respect. The rad6Δ1-99 mutant is defective in sporulation, shows reduced efficiency of DNA repair, but is proficient in UV mutagenesis. E3-dependent protein degradation by RAD6 could be essential for sporulation and could affect the efficiency of DNA repair.

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