Mitotic transcription and waves of gene reactivation during mitotic exit

Abstract

Although the genome is generally thought to be transcriptionally silent during mitosis, technical limitations have prevented sensitive mapping of transcription during mitosis and mitotic exit. Thus, the means by which the interphase expression pattern is transduced to daughter cells have been unclear. We used 5-ethynyluridine to pulse-label transcripts during mitosis and mitotic exit and found that many genes exhibit transcription during mitosis, as confirmed with fluorescein isothiocyanate-uridine 5'-triphosphate labeling, RNA fluorescence in situ hybridization, and quantitative reverse transcription polymerase chain reaction. The first round of transcription immediately after mitosis primarily activates genes involved in the growth and rebuilding of daughter cells, rather than cell type-specific functions. We propose that the cell's transcription pattern is largely retained at a low level through mitosis, whereas the amplitude of transcription observed in interphase is reestablished during mitotic exit.

Fig 1

EU-RNA-Seq and direct FITC-UTP labeling reveal extensive transcription in mitosis. (A) Pulse-labeling during mitosis and mitotic exit. (B) Reads span exons and introns, not intergenic regions; y-axis = FPM. (C) A representative transcript with an FPKM of 19. (D) FPKMs of mitotically-expressed transcripts, in mitosis and in async. bar = mean, whiskers = quartiles, p < 0.001, n=8,074. Interphase (E–G) or mitotic (H–M) cells labeled with FITC-UTP; white box magnified in (K–M). Interphase (N–P) or mitotic (Q–V) cells treated with α-amanitin and labeled with FITC-UTP; white box magnified in (Q–S), arrow = RNA signal, arrowhead = no RNA signal.

Fig 2

Markedly active genes in mitosis. Naturally-occurring mitotic cells in an async. population stained for DAPI (A, E, I) and exonic (B, F, J) and intronic (C, G, K) RNA. Colocalization at primary transcripts (D, H, L); white arrows = exon, yellow arrows = intron. (M) FPKMs of mitotically-enriched genes in mitotic and async. cells; bar = mean, whiskers = quartiles, p < 0.001, n=484. (N) Representative GO categories for mitotically-enriched genes.

Fig. 3

Transcription reactivation during mitotic exit. (A) Z-scores of transcripts (rows) that first increase ≥1.5 fold over mitosis, rank ordered within each time point (columns). (B) Liver genes during mitotic exit; y-axis = FPM, colored bars to the left indicate mitotic or mitotic exit stages generally observed at each time point.

Fig. 4

Basic cell functions are prioritized over cell specificity during mitotic exit. (A) Representative GO categories for 40–80′. (B) FPKMs of liver-specific genes; bar = mean, whiskers = quartiles, p < 0.001 from 300′ to async., n=149. (E) Increase in eRNA (29) within 100 kb of transcription start site (TSS) during mitotic exit; bar = mean, whiskers = quartiles, n.s = not significant, blue = comparison to mitosis, red = comparison to previous time point, *p<0.05, ***p<0.001.