p1-vv85, a new maize variegated pericarp allele, contains an insertion similar to the CACTA element, Mpi1 --Wang Y, Peterson, T The maize p1 gene encodes a Myb-like factor that regulates transcription of structural genes, including a1 and c2, required for red phlobaphene pigmentation in certain floral organs, including kernel, pericarp, and cob. We obtained a new p1 mutant allele designated p1-vv85, from Dr. Peter A. Peterson, ISU. The p1-vv85 allele specifies variegated pericarp and cob, and exhibits a very high rate of reversion to a state giving red pericarp and cob. Genetic analysis suggested that the mutant phenotype is not associated with Ac or En/Spm transposable element systems (P.A. Peterson, personal communication). Genomic Southern analysis indicated that p1-vv85 contains a 1.2 kb insertion in p1 gene intron 2, compared with its progenitor, P1-rr107B (from the Maize Genetics Stock Center). This region was amplified by PCR and found to contain a 1168bp insertion which has characteristics of the CACTA transposable element family: the insertion is bounded by 3 bp target site duplications, and it has 13bp TIR (terminal inverted repeats) which are identical to that of the CACTA transposable element Mpi1 (Weydemann et al., MNL 62: 48,1988). The insertion also has structured subterminal regions that contain 12 copies and 6 copies of a 12bp motif at the 5- and 3-ends of the fragment, respectively. Northern blot analysis revealed that the p1 gene transcript level is greatly reduced in p1-vv85 pericarp, while a revertant allele contains a normal level of p1 transcripts. Surprisingly, Southern blot analysis detected no difference in the structures of the p1-vv85 allele and two independent revertant alleles. Specifically, no evidence of excision of the 1168 bp insertion could be detected by either genomic Southern blot or PCR. Thus, the insertion in p1-vv85 may regulate p1 gene expression by an epigenetic mechanism. Analysis of DNA methylation of the p-vv85 mutant and a revertant allele revealed some differences in methylation of the insertion sequence, but not in the flanking p1 gene sequences. This result is consistent with a model in which methylation of the insertion sequence may affect the binding of a suppressor-like factor to the element. In summary, the p1-vv85 allele contains a CACTA element insertion relative to the progenitor P1-rr107B allele. However, the apparent "reversion" of p1-vv85 to a functional P1-rr state does not result from element excision, but rather by relief of suppression, which is correlated with the methylation state of the CACTA element.
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