Abstract
The centromere, in which kinetochore proteins are assembled, plays an important role in the accurate congression and segregation of chromosomes during cell mitosis. Although the function of the centromere and kinetochore is conserved from monocentric to holocentric, the DNA sequences of the centromere and components of the kinetochore are varied among different species. Given the lack of core centromere protein A (CENP-A) and CENP-C in the lepidopteran silkworm Bombyx mori, which possesses holocentric chromosomes, here we investigated the role of CENP-N, another important member of the centromere protein family essential for kinetochore assembly. For the first time, cellular localization and RNA interference against CENP-N have confirmed its kinetochore function in silkworms. To gain further insights into the regulation of CENP-N in the centromere, we analyzed the affinity-purified complex of CENP-N by mass spectrometry and identified 142 interacting proteins. Among these factors, we found that the chaperone protein heat shock cognate 70 (HSC70) is able to regulate the stability of CENP-N by prohibiting ubiquitin–proteasome pathway, indicating that HSC70 could control cell cycle-regulated degradation of CENP-N at centromeres. Altogether, the present work will provide a novel clue to understand the regulatory mechanism for the kinetochore activity of CENP-N during the cell cycle.
Highlights
The faithful transmission of genetic information among generations of cells requires the accurate congression and segregation of chromosomes during mitosis [1,2,3]
An important role in this process is performed by the centromere, a specialized chromatin region in the chromosome marked by a specific histone H3 variant CenH3, named centromere protein A (CENP-A), instead of the canonical histone H3 [4,5,6,7]
When we examined cell mitosis after CENP-N RNA interference (RNAi), it was clearly shown that knockdown of CENP-N significantly induced deficient congression and segregation of chromosomes at metaphase (Figure 1C)
Summary
The faithful transmission of genetic information among generations of cells requires the accurate congression and segregation of chromosomes during mitosis [1,2,3]. Based on the complete genomic information of the silkworm, previous work has identified several inner and outer kinetochore proteins, including eight putative CCAN proteins (CENP-E, -I, -K, -L, -M, -N, -S, and -X) and four KMN subunits (Ndc, Mis, Dsn, and Nnf1) [18,28,29]. The functions of these kinetochore proteins or unknown components in centromere formation, still need to be explored. We concluded that HSC70 plays a critical role in regulating the stability of kinetochore protein CENP-N in silkworms
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