In-depth sequence analysis of the tomato chromosome 12 centromeric region: identification of a large CAA block and characterization of pericentromere retrotranposons

Abstract

We sequenced a continuous 326-kb DNA stretch of a microscopically defined centromeric region of tomato chromosome 12. A total of 84% of the sequence (270 kb) was composed of a nested complex of repeat sequences including 27 retrotransposons, two transposable elements, three MITEs, two terminal repeat retrotransposons in miniature (TRIMs), ten unclassified repeats and three chloroplast DNA insertions. The retrotransposons were grouped into three families of Ty3-Gypsy type long terminal repeat (LTR) retrotransposons (PCRT1-PCRT3) and one LINE-like retrotransposon (PCRT4). High-resolution fluorescence in situ hybridization analyses on pachytene complements revealed that PCRT1a occurs on the pericentromere heterochromatin blocks. PCRT1 was the prevalent retrotransposon family occupying more than 60% of the 326-kb sequence with 19 members grouped into eight subfamilies (PCRT1a-PCRT1h) based on LTR sequence. The PCRT1a subfamily is a rapidly amplified element occupying tens of megabases. The other PCRT1 subfamilies (PCRT1b-PCRT1h) were highly degenerated and interrupted by insertions of other elements. The PCRT1 family shows identity with a previously identified tomato-specific repeat TGR2 and a CENP-B like sequence. A second previously described genomic repeat, TGR3, was identified as a part of the LTR sequence of an Athila-like PCRT2 element of which four copies were found in the 326-kb stretch. A large block of trinucleotide microsatellite (CAA)n occupies the centromere and large portions of the flanking pericentromere heterochromatin blocks of chromosome 12 and most of the other chromosomes. Five putative genes in the remaining 14% of the centromere region were identified, of which one is similar to a transcription regulator (ToCPL1) and a candidate jointless-2 gene.

FISH mapping and molecular organization of the major repetitive sequences of tomato

“…For comparison, the hybridization patterns of Cot-1, Cot-10, Cot-100 and of the previously published tandem (Figure 3a). This repetitive element shares 82% sequence identity with a novel putative Ty3-Gypsy type pericentromere retrotransposon of tomato and contains the LTR-region of the previously described PCRT1a element (Yang et al 2005), which is similar to the Jinling element (Wang et al 2006). The TGRIII repeat ( Figure 3b) and the tomato specific Ty1-copia-like retrotransposon (data not shown) were found in the pericentromere heterochromatin.…”

“…The TGRIII repeat ( Figure 3b) and the tomato specific Ty1-copia-like retrotransposon (data not shown) were found in the pericentromere heterochromatin. The 509 bp TGRIII element is similar to a part of the LTR sequence of the PCRT2 element, which is another novel tomato Ty3-Gypsy type pericentromere retrotransposon, showing homology to Athila in Arabidopsis (Yang et al 2005). Like TGRII, TGRIII was not detected in the NOR regions and, overall, TGRIII appeared to be less abundant than TGRII.…”

“…When FISH experiments were carried out with the entire BAC 57J04 as a probe (results not shown), strong signals on all tomato centromeres were observed as well as weak signals in some pericentromeres. Sequence analyses showed that this BAC for the larger part is composed of retrotransposon-like sequences of the TGRIV-type and of homologues of the pericentromere retrotransposons PCRTY1, Y2, Y3, and Y4 (Yang et al 2005). Next to a part with high similarity to the GYPSODE1 element (nt 2401Y7000), the TGRIV element also contains a unique part at its 5 ¶ end (nt 1Y2400) that shows no homology with any entry in the database but which could be an LTR of an ancestral TGRIVtype retrotransposon.…”

“…The TGRIV sequences were compared against the genome sequence database (http://www. ncbi.nlm.nih.gov/) and newly characterized tomato transposons (Yang et al 2005) using BLAST and PipMaker (Schwartz et al 2000, http://bio.cse.psu. edu/cgi-bin/pipmaker?advanced).…”

“…Vosman & Arens (1997) showed that these microsatellite motifs are frequently associated with one other repetitive element, which they termed U30 (cf. Ganal et al 1988, Yang et al 2005, Wang et al 2006.…”