A library of yeast genomic MCM1 binding sites contains genes involved in cell cycle control, cell wall and membrane structure, and metabolism.

Abstract

The Saccharomyces cerevisiae MCM1 protein, which is essential for viability, participates in both transcription activation and repression as well as DNA replication. However, neither the full network of genes at which MCM1 acts nor whether MCM1 itself mediates a regulatory response is known. Thus far, sites of MCM1 action have been identified by chance during analysis of particular genes. To identify a more complete set of genes on which MCM1 acts, we isolated a library of yeast genomic sequences to which MCM1 binds and then identified known genes within this library. Fragments of genomic DNA, bound to bacterially expressed MCM1 protein, were collected on a nitrocellulose filter, cloned, and analyzed. This selected library contains a large number of genes. As expected, it is enriched for strong MCM1 binding sites and contains cell-type-specific genes known to require MCM1. In addition, it also includes sequences upstream (or near the 5' end) of a number of identified yeast genes that have not yet been shown to be controlled by MCM1. These include genes whose products are involved in (i) the control of cell cycle progression (CLN3, CLB2, and FAR1), (ii) synthesis and maintenance of cell wall or cell membrane structures (PMA1, PIS1, DIT1,2, and GFA1), (iii) cellular metabolism (PCK1, MET2, and CCP1), and (iv) production of a secreted glycoprotein which is heat shock inducible (HSP150). The previously unidentified MCM1 binding site in the essential PMA1 gene is required for expression of a PMA1:lacZ fusion gene, providing evidence that one site is functionally important. We speculate that MCM1 coordinates decisions about cell cycle progression with changes in cell wall integrity and metabolic activity. The presence in the library of three genes involved in cell cycle progression reinforces the idea that one of the functions of MCM1 is indeed analogous to that of the mammalian serum response factor.