The CNA1 Histone of the CiliateTetrahymena thermophilaIs Essential for Chromosome Segregation in the Germline Micronucleus

Abstract

Ciliated protozoans present several features of chromosome segregation that are unique among eukaryotes, including their maintenance of two nuclei: a germline micronucleus, which undergoes conventional mitosis and meiosis, and a somatic macronucleus that divides by an amitotic process. To study ciliate chromosome segregation, we have identified the centromeric histone gene in the Tetrahymena thermophila genome (CNA1). CNA1p specifically localizes to peripheral centromeres in the micronucleus but is absent in the macronucleus during vegetative growth. During meiotic prophase of the micronucleus, when chromosomes are stretched to twice the length of the cell, CNA1p is found localized in punctate spots throughout the length of the chromosomes. As conjugation proceeds, CNA1p appears initially diffuse, but quickly reverts to discrete dots in those nuclei destined to become micronuclei, whereas it remains diffuse and is gradually lost in developing macronuclei. In progeny of germline CNA1 knockouts, we see no defects in macronuclear division or viability of the progeny cells immediately following the knockout. However, within a few divisions, progeny show abnormal mitotic segregation of their micronucleus, with most cells eventually losing their micronucleus entirely. This study reveals a strong dependence of the germline micronucleus on centromeric histones for proper chromosome segregation.

Centromeric Histone H3 Is Essential for Vegetative Cell Division and for DNA Elimination during Conjugation in Tetrahymena thermophila

“…Some of the results described here confirm or extend the studies reported by Cervantes et al (6), while others differ. We also studied additional properties of Cna1p.…”

“…Both our results and those of Cervantes et al (6) indicate that Cna1p is a chimeric histone with a H3-like histone fold domain, similar to the CenH3s of other organisms. The micronuclear-specific localization of Cna1p-GFP in small dots whose number correlates closely with the number of micronuclear chromosomes and whose behavior during mitosis is as expected for centromeres argues strongly that Cna1p is the centromeric H3 in Tetrahymena.…”

“…While the studies described here were in review, a report (6) describing some of the properties of Tetrahymena CNA1 was published. Some of the results described here confirm or extend the studies reported by Cervantes et al (6), while others differ.…”

“…In conjugating cells in which the CNA1 gene has been knocked out in micronuclei but not in the parental macronuclei, scan RNAs (scnRNAs) accumulate, histone H3 K9 methylation occurs, and the chromodomain protein Pdd1p appears in developing MACs, but the DNA elimination structures are not formed. In wild-type cells, Cna1p is detected in the DNA elimination structures using a specific antibody against Cna1p (␣-Cna1p [6]). These results indicate that Cna1p plays a critical role in later stages of DNA elimination, and centromere regions may function as the initiation sites for the formation of DNA elimination structures.…”

The Condensin Complex Is Essential for Amitotic Segregation of Bulk Chromosomes, but Not Nucleoli, in the Ciliate Tetrahymena thermophila

“…The macronucleus differs from other eukaryotic nuclei in several respects. It is derived from the micronucleus during conjugation through elaborate genomic rearrangement of the five germ line chromosomes into more than 200 chromosome fragments ranging in size from 20 to more than 3,000 kb (for a review, see reference 52) that have telomeres but no centromeres (7). During vegetative growth, the macronucleus divides by an amitotic mechanism that remains ill defined.…”