By R. Baserga, L. Kaczmarek, B. Calabretta, R. Battini, S. Ferrari (auth.), Professor Dr. Widmar Tanner, Professor Dr. Dieter Gallwitz (eds.)
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4. Screen for trans-acting regulatory mutants. A strain containing a single copy of an H2A-ZacZ fusion gene integrated at the TRTl locus was mutagenized with EMS. Surivors were replica plated to minimal media plates containing XGal and incubated at 23° and 36°C. Dark blue colonies were selected for analysis of cell cycle regulation. putative mutants were grown exponentially (-) or arrested in S-phase (+) with hydroxyurea (HU), and the levels of H2A-ZacZ RNA were measured by a Sl nuclease protection assay.
4 x 10- 8 M a-factor. The different dose required for agglutination and cell division arrest compared to that required for projection formation is teleologically sensible. At iowa-factor concentrations produced when MATa cells are in the same environment as MATa cells, the MATa cells arrest division in G1 and synthesize a surface agglutinin in preparation for conjugation. Upon physical interaction of a MATa cell with a MATa cell, agglutination takes place and now the close proximity of the MATa cell should result in a much higher concentration of a-factor.
The transformants were sterile as MATa cells but fertile as MATa cells, demonstrating that the STE2 gene is needed for fertility only in the MATa cell. In MATa cells this deletion eliminates all three a-factor inducible responses: agglutination, cell division arrest, and projection formation. This result demonstrates that the STE2 gene pro- 27 duct is needed for all three responses. This result does not prove that a single receptor is involved in all three responses, but it does indicate that if more than one receptor is involved, the STE2 gene product is likely to be a component of all receptors.