Thiopurine S-methyltransferase pharmacogenetics: Autophagy as a mechanism for variant allozyme degradation

Fang Li, Liewei Wang, Rebecca J. Burgess, Richard M. Weinshilboum

Research output: Contribution to journalArticle

21 Scopus citations

Abstract

Objective Thiopurine S-methyltransferase (TPMT)*3A is degraded much more rapidly than is the 'wild-type' enzyme through a ubiquitin-proteasome- dependent process. It also forms aggresomes, suggesting a possible dynamic balance between degradation and aggregation. We set out to identify genes encoding proteins participating in these processes. Methods Green fluorescent protein tagged TPMT*3A was expressed in a Saccharomyces cerevisiae gene deletion library, and flow cytometry was used to screen for cells with high fluorescence intensity, indicating the loss of a gene essential for TPMT*3A degradation. Results Twenty-four yeast genes were identified in functional categories that included ubiquitin-dependent protein degradation, vesicle trafficking, and vacuolar degradation. The presence of genes encoding proteins involved in vesicular transport and vacuolar degradation suggested a possible role in TPMT*3A degradation for autophagy - a process not previously identified as a pharmacogenomic mechanism. In support of that hypothesis, TPMT*3A aggregates increased dramatically in mutants for vacuolar protease and autophagy-related genes. Furthermore, TPMT*3A expression in human cells induced autophagy, and small interfering RNA-mediated knockdown of ATG7, an autophagy-related human protein, enhanced TPMT*3A aggregation but not that of TPMT*3C or wild-type TPMT, indicating that autophagy contributes to TPMT*3A degradation in mammalian cells. We also demonstrated that UBE2G2, the human homologue of the E2 ubiquitin-conjugating enzyme identified during the yeast genetic screen, was involved in TPMT*3A degradation in human cells. Conclusion These results indicate that autophagy should be considered among mechanisms responsible for the effects of pharmacogenetically significant polymorphisms that alter encoded amino acids.

Original languageEnglish (US)
Pages (from-to)1083-1094
Number of pages12
JournalPharmacogenetics and Genomics
Volume18
Issue number12
DOIs
StatePublished - Dec 1 2008
Externally publishedYes

Keywords

  • Autophagy
  • Protein aggregation
  • Protein degradation
  • Protein misfolding
  • Thiopurine S-methyltransferase
  • TPMT
  • TPMT *3A
  • TPMT*3C

ASJC Scopus subject areas

  • Molecular Medicine
  • Molecular Biology
  • Genetics
  • Genetics(clinical)

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