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The identification of cDNAs that affect the mitosis-to-interphase transition in Schizosaccharomyces pombe, including
1Verna and Marrs McLean Department of Biochemistry, Baylor College of Medicine, Houston, Texas 77030, USA.
Abstract:
Perturbations of the spi1p GTPase system in fission yeast, caused by mutation or overexpression of several regulatory proteins, result in a unique terminal phenotype that includes condensed chromosomes, a wide medial septum, and a fragmented nuclear envelope. To identify potential regulators or targets of the spi1p GTPase system, a screen for cDNAs whose overexpression results in this terminal phenotype was conducted, and seven clones that represent three genes, named med1, med2, and med3 (mitotic exit defect), were identified. Their genetic interaction with the spi1p GTPase system was established by showing that the spi1p guanine nucleotide exchange factor mutant pim1-d1ts was hypersensitive to their overexpression. med1 encodes a homologue of the human Ran-binding protein, RanBP1, and has been renamed sbp1 (spi1-binding protein). sbp1p binds to spi1p-GTP and costimulates the GTPase-activating protein (GAP)-catalyzed GTPase activity. Cells in which sbp1p is depleted or overproduced phenocopy cells in which the balance between spi1p-GTP and spi1p-GDP is perturbed by other means. Therefore, sbp1p mediates and/or regulates the essential functions of the spi1p GTPase system. med2 and med3 encode novel fission yeast proteins that, based on our phenotypic analyses, are likely to identify additional regulators or effectors of the spi1p GTPase system.
Insights
Researchers identified new regulators of the spi1p GTPase system in fission yeast. Overexpression of these genes, including sbp1, causes a unique cell division defect, revealing their crucial roles in regulating cell cycle progression.
Area of Science:
- Cell Biology
- Molecular Biology
- Genetics
Background:
- The spi1p GTPase system is crucial for cell cycle regulation in fission yeast.
- Perturbations in this system lead to a distinct terminal phenotype, including chromosomal abnormalities and defects in cell division.
- Identifying regulators and targets of spi1p is essential for understanding its functions.
Purpose of the Study:
- To identify novel regulators and targets of the spi1p GTPase system.
- To characterize the function of newly identified genes involved in cell division.
- To elucidate the role of sbp1 in mediating spi1p GTPase activity.
Main Methods:
- Conducted a screen for cDNAs that cause a specific terminal phenotype upon overexpression.
- Utilized genetic interaction analysis, including hypersensitivity assays with a pim1-d1ts mutant.
- Performed biochemical assays to analyze the interaction between sbp1p and spi1p-GTP.
Main Results:
- Identified three genes (med1, med2, med3) whose overexpression phenocopies spi1p system perturbations.
- Renamed med1 as sbp1 (spi1-binding protein), a homolog of human RanBP1.
- Demonstrated that sbp1p binds spi1p-GTP and enhances its GTPase activity, indicating its role as a mediator.
Conclusions:
- sbp1p is a key mediator and regulator of the essential functions of the spi1p GTPase system.
- med2 and med3 encode novel proteins likely involved in regulating the spi1p GTPase system.
- These findings provide new insights into the molecular mechanisms governing cell division and GTPase signaling.