Parallel analysis of tagged deletion mutants efficiently identifies genes involved in endoplasmic reticulum biogenesis.
Increased levels of HMG-CoA reductase induce cell type- and isozyme-specific proliferation of the endoplasmic reticulum. In yeast, the ER proliferations induced by Hmg1p consist of nuclear-associated stacks of smooth ER membranes known as karmellae. To identify genes required for karmellae assembly, we compared the composition of populations of homozygous diploid S. cerevisiae deletion mutants following 20 generations of growth with and without karmellae. Using an initial population of 1,557 deletion mutants, 120 potential mutants were identified as a result of three independent experiments. Each experiment produced a largely non-overlapping set of potential mutants, suggesting that differences in specific growth conditions could be used to maximize the comprehensiveness of similar parallel analysis screens. Only two genes, UBC7 and YAL011W, were identified in all three experiments. Subsequent analysis of individual mutant strains confirmed that each experiment was identifying valid mutations, based on the mutant's sensitivity to elevated HMG-CoA reductase and inability to assemble normal karmellae. The largest class of HMG-CoA reductase-sensitive mutations was a subset of genes that are involved in chromatin structure and transcriptional regulation, suggesting that karmellae assembly requires changes in transcription or that the presence of karmellae may interfere with normal transcriptional regulation.
Wright
Robin
R
University of Minnesota, Department of Genetics, Cell Biology and Development, 321 Church Street, 6-160 Jackson Hall, Minneapolis, MN 55455, USA. wrightr@umn.edu
Parrish
Mark L
ML
Cadera
Emily
E
Larson
Lynnelle
L
Matson
Clinton K
CK
Garrett-Engele
Philip
P
Armour
Chris
C
Lum
Pek Yee
PY
Shoemaker
Daniel D
DD
D004271:DNA, Fungal / Q000235:genetics / Q000378:metabolism; D004721:Endoplasmic Reticulum / Q000235:genetics* / Q000378:metabolism; D017353:Gene Deletion; D015966:Gene Expression Regulation, Fungal; D005800:Genes, Fungal / Q000235:genetics* / Q000502:physiology; D006903:Hydroxymethylglutaryl CoA Reductases / Q000235:genetics* / Q000378:metabolism; D016296:Mutagenesis; D020411:Oligonucleotide Array Sequence Analysis; D012441:Saccharomyces cerevisiae / Q000235:genetics* / Q000378:metabolism; D014170:Transformation, Genetic / Q000235:genetics / Q000502:physiology
D004271:DNA, Fungal; D006903:Hydroxymethylglutaryl CoA Reductases