cells (tables S18 and S19). Last, the last transcripts
to increase are involved in cell cycle and DNA
replication (tables S20 and S21), as the cells are
preparing for S phase. To determine when liver-specific genes first increase, we analyzed the
time at which the 149 liver-specific genes ( 27)
expressed in HUH7 cells—a number similar to
that seen in cultured hepatocytes ( 28)—first increases over mitosis. Although liver-specific genes
are expressed throughout mitotic exit, most are
reactivated at later stages (Fig. 4B and fig. S10A).
Thus, HUH7 cells initially activate genes required
for building daughter cells at the beginning of
mitotic exit and then later activate cell type–
Although the first transcripts to increase are
among the shortest (fig. S10B), the longest early
genes are still activated before the shortest late
genes (fig. S10C). Gene ontology (GO) analysis of
the longest genes to come up at 40 and 80 min
reveals basic cell functions (fig. S10, C and D), as
observed when considering all genes (Fig. 4A).
Analysis of the shortest liver-specific genes indicates that they increase at later time points (fig.
S10, E and F). Thus, the time of activation of gene
classes relates to their function and not primarily
to gene size.
We also assessed enhancer RNA (eRNA) dynamics as a surrogate for enhancer activity. We
curated all intergenic human enhancers ( 29) for
detectable eRNAs in asynchronous HUH7 cells
and found them significantly down-regulated in
mitosis (fig. S11A). The majority of eRNAs increased
at the early time points during mitotic release (fig.
S11B), as did genes (Fig. 3A). Curating for the enhancer subset within 100 kb of the nearest TSS,
we found that eRNAs first increase around the
same time as their putative targets (Fig. 4C).
Therefore, enhancer and putative target gene reactivation appear concordant during mitotic exit.
We applied a sensitive approach to measuring
the transcriptome during mitosis and mitotic exit.
We found extensive residual transcription in mitosis and waves of transcription reactivation during
exit. RNA polymerases have long been known to
be stable in chromatin, persisting during salt-washes of nuclei that cause loss of transcription
factors ( 30). Thus, a low level of transcribing
RNAP2 could contribute to the inheritance of a
cell’s specific transcriptome pattern through mitosis. Because deoxyribonuclease hypersensitivity
also persists at promoters in mitosis, whereas hypersensitivity at enhancers ( 9) and long-range interactions generally do not ( 8), we suggest that in
mitosis, the promoter and its gene create rudimentary mitotic expression units (MEUs). MEUs retain
residual activity and function along the general
constraints of genes in yeast, which lack enhancers and long-range interactions. The MEU model
posits that the transcription pattern is largely
retained through mitosis by MEUs, whereas the
amplitude of transcription observed in interphase
is reestablished during mitotic exit.
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We thank J. Lerner, G. Blobel, C. Hsiung, and M. Capelson for their
comments on the manuscript; training grant T32GM00812 to
K.C.P.; and NIH grants 2P20GM103629 (COBRE) to H.L. and
GM36477 to K.S.Z. All genomic data are accessible at GSE87476.
Materials and Methods
Figs. S1 to S11
Tables S1 to S24
References ( 31–35)
23 November 2016; resubmitted 30 May 2017
Accepted 1 September 2017
Published online 14 September 2017