Transcriptome analysis of the binucleate ciliate Tetrahymena thermophila with asynchronous nuclear cell cycles.

Tetrahymena thermophila harbors two functionally and physically distinct nuclei within a shared cytoplasm. During vegetative growth, the "cell cycles" of the diploid micronucleus and polyploid macronucleus are offset. Micronuclear S phase initiates just before cytokinesis and is completed in daughter cells before onset of macronuclear DNA replication. Mitotic micronuclear division occurs mid-cell cycle, while macronuclear amitosis is coupled to cell division. Here we report the first RNA-seq cell cycle analysis of a binucleated ciliated protozoan. RNA was isolated across 1.5 vegetative cell cycles, starting with a macronuclear G1 population synchronized by centrifugal elutriation. Using MetaCycle, 3244 of the 26,000+ predicted genes were shown to be cell cycle regulated. Proteins present in both nuclei exhibit a single mRNA peak that always precedes their macronuclear function. Nucleus-limited genes, including nucleoporins and importins, are expressed before their respective nucleus-specific role. Cyclin D and A/B gene family members exhibit different expression patterns that suggest nucleus-restricted roles. Periodically expressed genes cluster into seven cyclic patterns. Four clusters have known PANTHER gene ontology terms associated with G1/S and G2/M phase. We propose that these clusters encode known and novel factors that coordinate micro- and macronuclear-specific events such as mitosis, amitosis, DNA replication, and cell division.

Meiotic DNA breaks associated with recombination in S. pombe.

In the fission yeast Schizosaccharomyces pombe, we have detected prominent DNA breaks that appeared shortly after premeiotic DNA replication. These breaks, like meiotic recombination, required the products of the six rec genes tested. Prominent breaks were detected at widely separated sites, about 100-300 kb apart, equivalent to about 50-150 sites per genome or approximately the number of meiotic recombination events. Certain features of these breaks are similar to those in the distantly related yeast Saccharomyces cerevisiae, the only other organism in which meiotic DNA breaks have been reported. Other features, however, appear to be different. These results suggest that, although DNA breaks may be a general feature of meiotic recombination, the breaks in S. pombe may play a role different from those in S. cerevisiae.