Toward an Understanding of Satellite DNA Function in Crustacea

A G+C-rich satellite DNA (stDNA) in Gecarcinus lateralis with a repeat unit of 2.1 kbp makes up approximately 3% (16,000 copies/nucleus) of the land crab genome. We used sequences from conserved domains of the repeat unit to construct probes for exploring the possibility of the repeat units being transcribed into RNA, their distribution in genomic DNA, and their methylation status. A transcription product hybridizing with TRU-EXON, a region of the satellite bordered by possible exon splice junctions, was found 3′END, a 368 bp fragment at the 3′-end of the satellite, were also found in several other crustaceans. Transcription products that hybridized to AMPL-3, a probe from an amplified segment of the satellite, were not found in land crabs but were found in lobsters. In land crabs, several poly(A)+RNA transcripts that hybridized to 3′END were specific for DNA strand, tissue, and intermolt stage. The diversity of transcripts suggests that transcription of the satellite is related to the organization of the repeated DNA in the crab genome, and that some satellite domains may be transcribed because of their position relative to functional genes. This observation is supported by the additional observation that the satellite is differentially methylated. Repeats organized in tandem in the satellite are methylated, whereas those interspersed in main component DNA (mcDNA) are not. Because DNA methylation is a means of silencing transcription, our results suggest that satellite sequences that are transcribed are those interpersed in mcDNA. DNA methylation patterns are also tissue-specific; of the seven tissues examined (gill, claw muscle, body muscle, limb buds, midgut gland, testis, and ovary), gill stDNA is the least methylated. The observed differences inmethylation levels are not attributable to stage in the intermolt cycle or to sex. This is the first report indicating the presence of 5-methylcytosine (m5C) in a crustacean.