Nucleolar dynamics in cellular senescence

Abstract

The nucleolus is a subnuclear structure widely known as the site of ribosome biogenesis. Recently, additional functions of the nucleolus have been described, one of which is serving as a stress sensor. When the nucleolus senses stressors, it causes nucleolar stress characterised by changes to nucleolar area and function, along with signalling pathways including p53.

Although alterations in nucleolar area were identified under nucleolar stress, the dynamic of nucleolar area triggered by different senescence inducers remains elusive. Besides, gene expression associated with enlarged nucleoli is not well understood. Here, I show nucleolar dynamics in different types of cellular senescence including oncogene-induced senescence (OIS), secondary senescence, and replicative senescence (RS). I have used human diploid fibroblast that were subjected to immunofluorescence with fibrillarin antibodies, which were subsequently quantitated by automated image analysis. I found enlarged nucleolar area upon the induction of OIS and RS but were not found in secondary senescence.

Furthermore, nucleolar expansion was more significant in OIS than RS. In addition to nucleolar hypertrophy, changes in nuclear area and the enrichment of senescence-associated heterochromatin foci (SAHF) formation were identified along with enlarged nucleoli in OIS. Notably, SAHF-positive cells exhibited significantly enlarged nucleoli over SAHF-negative cells, suggesting alterations in nucleolar area may link with SAHF formation. Conversely, secondary senescence did not give rise to such nuclear alteration and chromosomal rearrangements. RS showed increased nuclear area, but no enrichment of SAHF formation in conjunction with nucleolar enlargement. The examination of candidate genes related with alterations in nucleoli in OIS and RS revealed distinct gene expression. For example, ribosomal protein L5 and L11 were overexpressed in OIS, yet they were suppressed in RS.

The results demonstrate nucleolar stress in response to different senescence inducers leads to unique nucleolar dynamics, changes to nuclear and heterochromatin structure, and altered gene expression.