SAF-A promotes origin licensing and replication fork progression to ensure robust DNA replication

Abstract

The organisation of chromatin is closely intertwined with biological activities of chromosome domains, including transcription and DNA replication status. Scaffold attachment factor A (SAF-A), also known as Heteronuclear Ribonucleoprotein Protein U (HNRNPU), contributes to the formation of open chromatin structure. Here we demonstrate that SAF-A promotes the normal progression of DNA replication, and enables resumption of replication after inhibition. We report that cells depleted for SAF-A show reduced origin licensing in G1 phase, and consequently reduced origin activation frequency in S phase. Replication forks also progress less consistently in cells depleted for SAF-A, contributing to reduced DNA synthesis rate. Single-cell replication timing analysis revealed two distinct effects of SAF-A depletion: first, the boundaries between early- and late-replicating domains become more blurred; and second, SAF-A depletion causes replication timing changes that tend to bring regions of discordant domain compartmentalisation and replication timing into concordance. Associated with these defects, SAF-A-depleted cells show elevated -H2AX formation and tend to enter quiescence. Overall we find that SAF-A protein promotes robust DNA replication to ensure continuing cell proliferation.

Deficiency of the Heterogeneous Nuclear Ribonucleoprotein U locus leads to delayed hindbrain neurogenesis

“…The results of our transcriptional profiling showed a role of HNRNPU in chromatin organization, in line with previous reports that demonstrated the function of HNRNPU in chromatin compaction, DNA synthesis, and chromosome folding during mitosis in different cell types and conditions (Connolly et al, 2022; Nozawa et al, 2017; Sharp et al, 2020). Therefore, we sought to investigate whether HNRNPU deficiency affects chromatin organization in our human neural cell model.…”

“…Here, we provide novel evidence of the molecular and cellular consequences of HNRNPU deficiency in human neuronal cells, demonstrating that adequate levels of HNRNPU are needed in the early developmental transition from neural progenitors to developing neurons for proper neurogenesis. Our results, consistent with the earlier reports, show that HNRNPU expression is highest at early neural stem cell and progenitor stages and in a subpopulation of neural progenitors at the later stage of neuronal differentiation (Connolly et al, 2022; Sapir et al, 2022). Despite this high expression, results indicate that HNRNPU deficiency does not affect neural cells at this early stage.…”

“…For instance, HNRNPU modulates chromatin compaction by promoting chromatin accessibility in a dynamic yet structured fashion, dependent on its oligomerization status (Nozawa et al, 2017). Recent studies have also shown the involvement of HNRNPU in mitosis and cell division by changing its interactions with condensed chromatin and influencing DNA replication and sister chromatid separation (Connolly et al, 2022; Ma et al, 2011; Sharp et al, 2020). Several studies have reported a role for HNRNPU in splicing, promoting both exon inclusion and exclusion (Huelga et al, 2012; Jones et al, 2022; Ye et al, 2015).…”

Myeloid leukemia factor 1: A “double-edged sword” in health and disease

“…Manp localizes exclusively in the nucleus and redirects MLF1 into the nucleus ( 8 ). Recent studies have suggested that hnRNP-U regulates DNA replication, organizes large-scale chromosome structures, and protects the genome from instability ( 39 – 41 ). The effects of MLF on DNA synthesis have been previously discussed ( 19 ).…”

HNRNPU's multi‐tasking is essential for proper cortical development

“…Knockdown of the protein led to increased spacing between replication origins, slowing the replication fork progression, creating replication stress, and increasing cellular quiescence. [ 47 ] Shortly after exposure to ionizing irradiation and induction of double strands breaks, HNRNPU/SAF‐A transiently clusters at damage sites. This localization is regulated by phosphorylation at Ser59 by (DNA‐dependent protein kinase) DNA‐PK, thus suppressing base excision repair (BER) initiation and allowing the predominant nonhomologous end joining (NHEJ) repair to proceed.…”