YEAST VOL. 11: 1223-1231 (1995) Molecular Characterization of thezyxwvutsrqponmlkjihgfedcbaZYXWVUTSRQPONMLKJIHGFEDCBA PEL1 Gene Encoding a Putative Phosphatidylserine Synthase MARTIN JANITOR. ERNST JAROSCHi\F J. SCHWEYENt AND JULIUS SUBIK* Comenius University, Department oj Microbiology and Virology, 842 15 Bratislava, Slovak Republic ^Vienna Biocenter, Institute of Microbiology and Genetics, 1030 Wien, Austria Received 24 February 1995; accepted 6 June 1995 In the yeast Saccharomyces cerevisiae the PEL1 gene is essential for the viability of rho ~ Irho" petite mutants, and its mutation in respiring cells results in a pleiotropic phenotype. Results of complementation analysis with different subclones of chromosomal DNA and re-sequencing of the YCL4w-YCL3w segment of chromosome III demonstrate that the coding region of the PEL1 gene corresponds to 1467 bp. The size of the PEL1 transcript in Northern blot analysis was estimated to be approximately 1-5 kb. Transcription initiation in wild-type cells was found to occur at the position - 9 relative to the ATG. The PEL1 gene was moderately expressed irrespective of the state of the mitochondrial genome and the nature of the carbon sources. Disruption of the PEL1 gene was not lethal and resulted in the same phenotype as observed with the pell mutant, i.e. the cells were not able to survive ethidium bromide mutagenesis, were thermosensitive for growth on glucose at 37°C and failed to grow on minimal glycerol medium. Although the Pell protein exhibits significant similarity to a family of phosphatidylserine synthases, the disrupted PEL1 gene was not complemented by the multicopy plasmid-borne CHOI gene encoding an essential yeast phosphatidylserine synthase. The Accession Number of the PEL1 gene in the EMBL data library is Z48162. KEY WORDS — PEL1 gene; petite viability; phospholipid synthesis; yeast; Saccharomyces cerevisiae INTRODUCTION Saccharomyces cerevisiae is a />e?/fe-positive yeast species that can survive without mitochondrial DNA and hence without oxidative phosphoryla- tion (Dujon, 1981). In spite of this, its cells still need mitochondria and intramitochondrial ATP for reasons other than a supply of energy for growth (Subik et ah, 1972; Gbelska et ah, 1983; Baker and Schatz, 1981). This yeast is, there- fore, an ideal system for the genetic approach of studying the nucleo-mitochondrial interactions in eukaryotes. In the past decades, several yeast nuclear mutants have been described that were unable to survive the superimposition of the cyto- plasmic petite mutations over the nuclear one. Such petite-negative mutants of S. cerevisiae are represented by the mutant strain opllpet9, deficient in mitochondrial ATP/ADP translocation (Kovacova et ah, 1968); by the mutant pell, par- tially deficient in the biogenesis of mitochondria (Subik, 1974); by the mutant add, deficient in ""Corresponding author. CCC 0749-503X/95/131223-09 © 1995 by John Wiley & Sons Ltd constitutive alcohol dehydrogenase (Ciriacy, 1976) and by the mutant ymel, exhibiting an increased escape of DNA from mitochondria to the nucleus (Thorsness et ah, 1993). The pell mutant was found to exhibit a jpleio- tropic phenotype (Subik, 1974; Janitor and Subik, 1993). Its cells have an increased doubling time, are unable to grow on synthetic glycerol or ethanol medium and fail to grow on glucose medium at 37°C. The corresponding PEL1 gene has been mapped on chromosome I I I and isolated by complementation of the pell mutation. On the basis of the results of restriction analysis, partial sequence analysis and complementation activity of the subclones, the YCL4w open reading frame (ORF) has been found to co-localize with the PEL1 gene (Janitor et ah, 1992; Janitor and Subik, 1993). Recently, the computer re-examination of the published nucleotide sequence of chromo- some III indicated that the adjacent YCL3w ORF also belongs to the PEL1 gene, which likely en- codes a phosphatidylserine synthase (Koonin et ah, 1994).