Abstract
DNA replication occurs on mammalian chromosomes in a cell-type distinctive temporal order known as the replication timing program. We previously found that disruption of the noncanonical lncRNA genes ASAR6 and ASAR15 results in delayed replication timing and delayed mitotic chromosome condensation of human chromosomes 6 and 15, respectively. ASAR6 and ASAR15 display random monoallelic expression and display asynchronous replication between alleles that is coordinated with other random monoallelic genes on their respective chromosomes. Disruption of the expressed allele, but not the silent allele, of ASAR6 leads to delayed replication, activation of the previously silent alleles of linked monoallelic genes, and structural instability of human chromosome 6. In this report, we describe a second lncRNA gene (ASAR6-141) on human chromosome 6 that when disrupted results in delayed replication timing in cis. ASAR6-141 is subject to random monoallelic expression and asynchronous replication and is expressed from the opposite chromosome 6 homolog as ASAR6. ASAR6-141 RNA, like ASAR6 and ASAR15 RNAs, contains a high L1 content and remains associated with the chromosome territory where it is transcribed. Three classes of cis-acting elements control proper chromosome function in mammals: origins of replication, centromeres, and telomeres, which are responsible for replication, segregation, and stability of all chromosomes. Our work supports a fourth type of essential chromosomal element, the “Inactivation/Stability Center,” which expresses ASAR lncRNAs responsible for proper replication timing, monoallelic expression, and structural stability of each chromosome.
Highlights
Numerous reports over the past 50+ yr have described an abnormal DNA replication phenotype affecting individual chromosomes in mitotic preparations from mammalian cells
HTD114 cells are a human fibrosarcoma cell line, where we previously carried out the Cre/loxP screen that led to the identification and functional characterization of Autosomal RNA on chromosome 6 (ASAR6) and ASAR15 (Breger et al 2005; Stoffregen et al 2011; Donley et al 2015)
We found that vlinc273 and KCNQ5 show similar levels of monoallelic expression with single sites of RNA hybridization in ≥84% of cells (Fig. 3A–E; Supplemental Fig. S2C)
Summary
Numerous reports over the past 50+ yr have described an abnormal DNA replication phenotype affecting individual chromosomes in mitotic preparations from mammalian cells (for review, see Thayer 2012). Type further, we developed a Cre/loxP system that allowed us to create chromosome translocations in a precise and controllable manner (Breger et al 2004, 2005). Using this Cre/loxP system, we carried out a screen in human cells designed to identify loxP integration sites that generate translocated chromosomes with DRT/DMC (Breger et al 2004, 2005; Stoffregen et al 2011; Donley et al 2015). ∼5% of translocations induced by two different mechanisms (IR or Cre/loxP) result in DRT/DMC
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