Analysis of a High-Throughput Yeast Two-Hybrid System and Its Use to Predict the Function of Intracellular Proteins Encoded Within the Human MHC Class III Region

Genomics. 2004 Jan;83(1):153-67. doi: 10.1016/s0888-7543(03)00235-0.

Abstract

High-throughput (HTP) protein-interaction assays, such as the yeast two-hybrid (Y2H) system, are enormously useful in predicting the functions of novel gene-products. HTP-Y2H screens typically do not include all of the reconfirmation and specificity tests used in small-scale studies, but the effects of omitting these steps have not been assessed. We performed HTP-Y2H screens that included all standard controls, using the predicted intracellular proteins expressed from the human MHC class III region, a region of the genome associated with many autoimmune diseases. The 91 novel interactions identified provide insight into the potential functions of many MHC genes, including C6orf47, LSM2, NELF-E (RDBP), DOM3Z, STK19, PBX2, RNF5, UAP56 (BAT1), ATP6G2, LST1/f, BAT2, Scythe (BAT3), CSNK2B, BAT5, and CLIC1. Surprisingly, our results predict that 1/3 of the proteins may have a role in mRNA processing, which suggests clustering of functionally related genes within the human genome. Most importantly, our analysis shows that omitting standard controls in HTP-Y2H screens could significantly compromise data quality.

MeSH terms

  • Genes, Reporter / genetics
  • Humans
  • K562 Cells
  • Major Histocompatibility Complex / genetics*
  • Pilot Projects
  • Protein Binding
  • Proteins / genetics
  • Proteins / metabolism*
  • RNA, Messenger / metabolism
  • Recombinant Fusion Proteins / genetics
  • Recombinant Fusion Proteins / metabolism
  • Saccharomyces cerevisiae / genetics
  • Signal Transduction
  • Transcription Factors / genetics
  • Transcription Factors / metabolism
  • Two-Hybrid System Techniques*
  • Ubiquitin / metabolism

Substances

  • Proteins
  • RNA, Messenger
  • Recombinant Fusion Proteins
  • Transcription Factors
  • Ubiquitin