General Assessment
This study makes a convincing case that centromeric noncoding RNAs play an important role in chromosomal integrity, and also provides an RNA transcript set that will facilitate future studies of centromere transcription. Given the difficulty of targeting highly repetitive centromeric sequence, the presented data and interpretations make impressive demonstrations on cenRNA localization and function. In particular, the in vivo immunofluorescence experiments clearly demonstrate that an individual cenRNA (cenRNA#4) localizes near centromeres. If the authors choose to make an addition to the study, they may consider adding an IF image of cells experiencing chromosomal segregation defects, as it would help illustrate the direct observations to readers who are unfamiliar with the field. In addition, future readers with backgrounds in genomics will likely be interested in knowing the defining feature of each of the four identified cenRNA classes, and whether cenRNA loci are generally near canonical promoters and/or polyadenylation signals. Overall, this work contributes exciting findings in support of a fascinating model that explains how centromeric RNAs target and load CENP-A protein to the centromere, ensuring proper chromosomal segregation during mitosis.