An aberrant immune-epithelial progenitor niche drives viral lung sequelae.

The long-term physiological consequences of respiratory viral infections, particularly in the aftermath of the COVID-19 pandemic-termed post-acute sequelae of SARS-CoV-2 (PASC)-are rapidly evolving into a major public health concern. While the cellular and molecular aetiologies of these sequelae are poorly defined, increasing evidence implicates abnormal immune responses and/or impaired organ recovery after infection. However, the precise mechanisms that link these processes in the context of PASC remain unclear.

Repair of noise-induced damage to stereocilia F-actin cores is facilitated by XIRP2 and its novel mechanosensor domain.

Prolonged exposure to loud noise has been shown to affect inner ear sensory hair cells in a variety of deleterious manners, including damaging the stereocilia core. The damaged sites can be visualized as 'gaps' in phalloidin staining of F-actin, and the enrichment of monomeric actin at these sites, along with an actin nucleator and crosslinker, suggests that localized remodeling occurs to repair the broken filaments. Herein, we show that gaps in mouse auditory hair cells are largely repaired within 1 week of traumatic noise exposure through the incorporation of newly synthesized actin.

Stable maintenance of the Mre11-Rad50-Nbs1 complex is sufficient to restore the DNA double-strand break response in cells lacking RecQL4 helicase activity.

The proper cellular response to DNA double-strand breaks (DSBs) is critical for maintaining the integrity of the genome. RecQL4, a DNA helicase of which mutations are associated with Rothmund-Thomson syndrome (RTS), is required for the DNA DSB response. However, the mechanism by which RecQL4 performs these essential roles in the DSB response remains unknown.

RecQL4 tethering on the pre-replicative complex induces unscheduled origin activation and replication stress in human cells.

Sequential activation of DNA replication origins is precisely programmed and critical to maintaining genome stability. RecQL4, a member of the conserved RecQ family of helicases, plays an essential role in the initiation of DNA replication in mammalian cells. Here, we showed that RecQL4 protein tethered on the pre-replicative complex (pre-RC) induces early activation of late replicating origins during S phase.

ATM activation is impaired in human cells defective in RecQL4 helicase activity.

RecQL4 has been shown to be involved in DNA replication and repair, but its role in DNA damage checkpoint pathway has not been reported. Here, we show that RecQL4 plays an important role in the activation of ataxia telangiectasia mutated (ATM)-dependent checkpoint pathway in human cells. Cells depleted with RecQL4 or Rothmund-Thomson syndrome cells showed significant impairment in the activation of ATM and the downstream effector proteins such as checkpoint kinase 2 and p53 after DNA damage.

ASF1a Promotes Non-homologous End Joining Repair by Facilitating Phosphorylation of MDC1 by ATM at Double-Strand Breaks.

Double-strand breaks (DSBs) of DNA in eukaryotic cells are predominantly repaired by non-homologous end joining (NHEJ). The histone chaperone anti-silencing factor 1a (ASF1a) interacts with MDC1 and is recruited to sites of DSBs to facilitate the interaction of phospho-ATM with MDC1 and phosphorylation of MDC1, which are required for the recruitment of RNF8/RNF168 histone ubiquitin ligases. Thus, ASF1a deficiency reduces histone ubiquitination at DSBs, decreasing the recruitment of 53BP1, and decreases NHEJ, rendering cells more sensitive to DSBs.

MCM8-9 complex promotes resection of double-strand break ends by MRE11-RAD50-NBS1 complex.

MCM8-9 complex is required for homologous recombination (HR)-mediated repair of double-strand breaks (DSBs). Here we report that MCM8-9 is required for DNA resection by MRN (MRE11-RAD50-NBS1) at DSBs to generate ssDNA. MCM8-9 interacts with MRN and is required for the nuclease activity and stable association of MRN with DSBs.

RecQL4 is required for the association of Mcm10 and Ctf4 with replication origins in human cells.

Though RecQL4 was shown to be essential for the initiation of DNA replication in mammalian cells, its role in initiation is poorly understood. Here, we show that RecQL4 is required for the origin binding of Mcm10 and Ctf4, and their physical interactions and association with replication origins are controlled by the concerted action of both CDK and DDK activities. Although RecQL4-dependent binding of Mcm10 and Ctf4 to chromatin can occur in the absence of pre-replicative complex, their association with replication origins requires the presence of the pre-replicative complex and CDK and DDK activities.

ATR checkpoint kinase and CRL1βTRCP collaborate to degrade ASF1a and thus repress genes overlapping with clusters of stalled replication forks.

Many agents used for chemotherapy, such as doxorubicin, interfere with DNA replication, but the effect of this interference on transcription is largely unknown. Here we show that doxorubicin induces the firing of dense clusters of neoreplication origins that lead to clusters of stalled replication forks in gene-rich parts of the genome, particularly on expressed genes. Genes that overlap with these clusters of stalled forks are actively dechromatinized, unwound, and repressed by an ATR-dependent checkpoint pathway.

Mimosine arrests the cell cycle prior to the onset of DNA replication by preventing the binding of human Ctf4/And-1 to chromatin via Hif-1α activation in HeLa cells.

Though the G(1) checkpoint in mammalian cells has been known for decades, the molecular targets that prevent S-phase entry remain unknown. Mimosine is a rare plant amino acid that arrests the cell cycle in the G(1) phase before entry into S phase. Here, we show that mimosine interrupts the binding of Ctf4 to chromatin, which is essential for the initiation of DNA replication in HeLa cells, and this effect is mediated by the Hif-1α-dependent increase in the level of p27.

Per3, a circadian gene, is required for Chk2 activation in human cells.

PER3 is a member of the PERIOD genes, but does not play essential roles in the circadian clock. Depletion of Per3 by siRNA almost completely abolished activation of checkpoint kinase 2 (Chk2) after inducing DNA damage in human cells. In addition, Per3 physically interacted with ATM and Chk2.

Assembly of the Cdc45-Mcm2-7-GINS complex in human cells requires the Ctf4/And-1, RecQL4, and Mcm10 proteins.

In eukaryotes, the activation of the prereplicative complex and assembly of an active DNA unwinding complex are critical but poorly understood steps required for the initiation of DNA replication. In this report, we have used bimolecular fluorescence complementation assays in HeLa cells to examine the interactions between Cdc45, Mcm2-7, and the GINS complex (collectively called the CMG complex), which seem to play a key role in the formation and progression of replication forks. Interactions between the CMG components were observed only after the G(1)/S transition of the cell cycle and were abolished by treatment of cells with either a CDK inhibitor or siRNA against the Cdc7 kinase.

ATR-dependent activation of p38 MAP kinase is responsible for apoptotic cell death in cells depleted of Cdc7.

Cdc7 is a serine/threonine kinase that plays essential roles in the initiation of eukaryotic DNA replication and checkpoint response. In previous studies, depletion of Cdc7 by small interfering RNA was shown to induce an abortive S phase that led to the cell cycle arrest in normal human fibroblasts and apoptotic cell death in various cancer cells. Here we report that stress-activated p38 MAP kinase was activated and responsible for apoptotic cell death in Cdc7-depleted HeLa cells.

Senescence-associated beta-galactosidase is lysosomal beta-galactosidase.

Replicative senescence limits the proliferation of somatic cells passaged in culture and may reflect cellular aging in vivo. The most widely used biomarker for senescent and aging cells is senescence-associated beta-galactosidase (SA-beta-gal), which is defined as beta-galactosidase activity detectable at pH 6.0 in senescent cells, but the origin of SA-beta-gal and its cellular roles in senescence are not known.

Induction of senescence-like state and suppression of telomerase activity through inhibition of HPV E6/E7 gene expression in cells immortalized by HPV16 DNA.

The E6 and E7 oncoproteins of human papillomavirus (HPV) play a major role in the development of cervical carcinoma. In this study, a recombinant adenovirus that expresses the bovine papillomavirus (BPV) E2, which has been shown to inhibit HPV early gene expression, was delivered to two HPV-immortalized cell lines as well as CaSki, a cervical carcinoma cell line. We tested whether the carcinoma and the immortal cells were equally affected by the expression of BPV E2.