PI-SceI (VDE), a homing endonuclease with protein splicing activity, is a genomic parasite in the VMA1 gene of Saccharomyces cerevisiae. In a heterozygous diploid of the VDE-less VMA1 allele and a VDE-containing VMA1 allele, VDE specifically cleaves its recognition sequence (VRS) in the VDE-less VMA1 allele at meiosis, followed by 'homing', i.e. a conversion to a VDE-containing allele. We found that upon VDE expression, homing of a marker gene at an extragenic locus occurs only when a 45 bp element containing the VRS is inserted at its allelic site, while mutants of VDE with no endonuclease activity lack authentic extragenic homing activity. Thus, both the VRS and VDE are required for homing. Insertion of the VRS in a homozygous diploid significantly lowered the spore germination ability, indicating that a template for gene repair at its allelic locus is essential for efficient homing and survival of yeast cells.

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VMA1-derived endonuclease (VDE), a site-specific endonuclease in Saccharomyces cerevisiae, enters the nucleus to generate a double-strand break in the VDE-negative allelic locus, mediating the self-propagating gene conversion called homing. Although VDE is excluded from the nucleus in mitotic cells, it relocalizes at premeiosis, becoming localized in both the nucleus and the cytoplasm in meiosis. The nuclear localization of VDE is induced by inactivation of TOR kinases, which constitute central regulators of cell differentiation in S. cerevisiae, and by nutrient depletion. A functional genomic approach revealed that at least two karyopherins, Srp1p and Kap142p, are required for the nuclear localization pattern. Genetic and physical interactions between Srp1p and VDE imply direct involvement of karyopherin-mediated nuclear transport in this process. Inactivation of TOR signaling or acquisition of an extra nuclear localization signal in the VDE coding region leads to artificial nuclear localization of VDE and thereby induces homing even during mitosis. These results serve as evidence that VDE utilizes the host systems of nutrient signal transduction and nucleocytoplasmic transport to ensure the propagation of its coding region.Homing endonucleases are site-specific endonucleases encoded by introns and inteins (internal protein sequences) to promote homing of their genetic elements (homing endonuclease genes [HEG]) into allelic intronless and inteinless alleles, respectively (26). Homing endonuclease genes, widely found in Archaea, Bacteria, and Eukarya, are thought to be parasitic mobile genetic elements because they have no known host functions and their catalytic activity results in self-propagation.In Saccharomyces cerevisiae, the VMA1 intein encodes a homing endonuclease (14) termed the VMA1-derived endonuclease (VDE) (also called PI-SceI [36]), which is produced by an autocatalytic protein splicing reaction (19,25). When heterozygous VMA1(ϩ)/VMA1(⌬) diploid cells undergo meiosis under nutrient-deficient conditions, VDE expressed from the intein-containing allele [referred to as VMA1(ϩ)] specifically cleaves the VDE recognition sequence in the inteinless VMA1 allele [referred to as VMA1(⌬)] in the nuclear genome (4, 14) to make a double-strand break. The double-strand break is then repaired, with the VMA1(ϩ) allele as a template. This leads to the unidirectional gene conversion of VMA1(⌬) to VMA1(ϩ), which is called homing. Consequently, the VDE coding region exhibits super-Mendelian inheritance, allowing it to spread in yeast populations even though VDE confers no known benefit to the host yeast cells.VDE-mediated homing has the following two characteristic features: VDE homing occurs only in meiosis and not in mitosis even if the VMA1(ϩ) and VMA1(⌬) alleles coexist (14). In contrast, other homing endonucleases initiate homing in vitro and in vivo once the HEG(ϩ) allele is accompanied by the HEG(Ϫ) allele (7,29). Meiosis is assumed to be the best time for homing because of the homologue pairing and bias towar...

Most tumours are thought to arise through oncogenic cell generation followed by additional mutations. How a new oncogenic cell primes tumorigenesis by acquiring additional mutations remains unclear. We show that an additional TP53 mutation stimulates primary tumorigenesis by switching oncogene-induced senescence from a tumour suppressor to a driver. Zebrafish imaging reveals that a newly emerged oncogenic cell with the RasG12V mutation becomes senescent and is eliminated from the epithelia, which is prevented by adding a TP53 gain-of-function mutation (TP53R175H) into RasG12V cells. Surviving RasG12V-TP53R175H double-mutant cells senesce and secrete senescence-associated secretory phenotype (SASP)-related inflammatory molecules that convert neighbouring normal cells into SASP factor-secreting senescent cells, generating a heterogeneous tumour-like cell mass. We identify oncogenic cell behaviours that may control the initial human tumorigenesis step. Ras and TP53 mutations and cellular senescence are frequently detected in human tumours; similar switching may occur during the initial step of human tumorigenesis.

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