Inactive Parp2 causes Tp53-dependent lethal anemia by blocking replication-associated nick ligation in erythroblasts.

PARP1&2 enzymatic inhibitors (PARPi) are promising cancer treatments. But recently, their use has been hindered by unexplained severe anemia and treatment-related leukemia. In addition to enzymatic inhibition, PARPi also trap PARP1&2 at DNA lesions. Here, we report that unlike Parp2 -/- mice, which develop normally, mice expressing catalytically-inactive Parp2 (E534A, Parp2 EA/EA ) succumb to Tp53- and Chk2 -dependent erythropoietic failure in utero , mirroring Lig1 -/- mice. While DNA damage mainly activates PARP1, we demonstrate that DNA replication activates PARP2 robustly. PARP2 is selectively recruited and activated by 5'-phosphorylated nicks (5'p-nicks) between Okazaki fragments, typically resolved by Lig1. Inactive PARP2, but not its active form or absence, impedes Lig1- and Lig3-mediated ligation, causing dose-dependent replication fork collapse, particularly harmful to erythroblasts with ultra-fast forks. This PARylation-dependent structural function of PARP2 at 5'p-nicks explains the detrimental effects of PARP2 inhibition on erythropoiesis, revealing the mechanism behind the PARPi-induced anemia and leukemia, especially those with TP53/CHK2 loss. Significance: This work shows that the hematological toxicities associated with PARP inhibitors stem not from impaired PARP1 or PARP2 enzymatic activity but rather from the presence of inactive PARP2 protein. Mechanistically, these toxicities reflect a unique role of PARP2 at 5'-phosphorylated DNA nicks during DNA replication in erythroblasts.

Chromatin architecture at susceptible gene loci in cerebellar Purkinje cells characterizes DNA damage-induced neurodegeneration.

The pathogenesis of inherited genome instability neurodegenerative syndromes remains largely unknown. Here, we report new disease-relevant murine models of genome instability­driven neurodegeneration involving disabled ATM and APTX that develop debilitating ataxia. We show that neurodegeneration and ataxia result from transcriptional interference in the cerebellum via aberrant messenger RNA splicing. Unexpectedly, these splicing defects were restricted to only Purkinje cells, disrupting the expression of critical homeostatic regulators including ITPR1, GRID2, and CA8. Abundant genotoxic R loops were also found at these Purkinje cell gene loci, further exacerbating DNA damage and transcriptional disruption. Using ATAC-seq to profile global chromatin accessibility in the cerebellum, we found a notably unique chromatin conformation specifically in Purkinje chromatin at the affected gene loci, thereby promoting susceptibility to DNA damage. These data reveal the pathogenic basis of DNA damage in the nervous system and suggest chromatin conformation as a feature in directing genome instability­associated neuropathology.

Genome instability independent of type I interferon signaling drives neuropathology caused by impaired ribonucleotide excision repair.

Aicardi-Goutières syndrome (AGS) is a monogenic type I interferonopathy characterized by neurodevelopmental defects and upregulation of type I interferon signaling and neuroinflammation. Mutations in genes that function in nucleic acid metabolism, including RNASEH2, are linked to AGS. Ribonuclease H2 (RNASEH2) is a genome surveillance factor critical for DNA integrity by removing ribonucleotides incorporated into replicating DNA. Here we show that RNASEH2 is necessary for neurogenesis and to avoid activation of interferon-responsive genes and neuroinflammation. Cerebellar defects after RNASEH2B inactivation are rescued by p53 but not cGAS deletion, suggesting that DNA damage signaling, not neuroinflammation, accounts for neuropathology. Coincident inactivation of Atm and Rnaseh2 further affected cerebellar development causing ataxia, which was dependent upon aberrant activation of non-homologous end-joining (NHEJ). The loss of ATM also markedly exacerbates cGAS-dependent type I interferon signaling. Thus, DNA damage-dependent signaling rather than type I interferon signaling underlies neurodegeneration in this class of neurodevelopmental/neuroinflammatory disease.

Histone H3.3 K27M Accelerates Spontaneous Brainstem Glioma and Drives Restricted Changes in Bivalent Gene Expression.

Diffuse intrinsic pontine gliomas (DIPGs) are incurable childhood brainstem tumors with frequent histone H3 K27M mutations and recurrent alterations in PDGFRA and TP53. We generated genetically engineered inducible mice and showed that H3.3 K27M enhanced neural stem cell self-renewal while preserving regional identity. Neonatal induction of H3.3 K27M cooperated with activating platelet-derived growth factor receptor α (PDGFRα) mutant and Trp53 loss to accelerate development of diffuse brainstem gliomas that recapitulated human DIPG gene expression signatures and showed global changes in H3K27 posttranslational modifications, but relatively restricted gene expression changes. Genes upregulated in H3.3 K27M tumors were enriched for those associated with neural development where H3K27me3 loss released the poised state of apparently bivalent promoters, whereas downregulated genes were enriched for those encoding homeodomain transcription factors.

Apurinic endonuclease-1 preserves neural genome integrity to maintain homeostasis and thermoregulation and prevent brain tumors.

Frequent oxidative modification of the neural genome is a by-product of the high oxygen consumption of the nervous system. Rapid correction of oxidative DNA lesions is essential, as genome stability is a paramount determinant of neural homeostasis. Apurinic/apyrimidinic endonuclease 1 (APE1; also known as "APEX1" or "REF1") is a key enzyme for the repair of oxidative DNA damage, although the specific role(s) for this enzyme in the development and maintenance of the nervous system is largely unknown. Here, using conditional inactivation of murine Ape1, we identify critical roles for this protein in the brain selectively after birth, coinciding with tissue oxygenation shifting from a placental supply to respiration. While mice lacking APE1 throughout neurogenesis were viable with little discernible phenotype at birth, rapid and pronounced brain-wide degenerative changes associated with DNA damage were observed immediately after birth leading to early death. Unexpectedly, Ape1Nes-cre mice appeared hypothermic with persistent shivering associated with the loss of thermoregulatory serotonergic neurons. We found that APE1 is critical for the selective regulation of Fos1-induced hippocampal immediate early gene expression. Finally, loss of APE1 in combination with p53 inactivation resulted in a profound susceptibility to brain tumors, including medulloblastoma and glioblastoma, implicating oxidative DNA lesions as an etiologic agent in these diseases. Our study reveals APE1 as a major suppressor of deleterious oxidative DNA damage and uncovers specific and broad pathogenic consequences of respiratory oxygenation in the postnatal nervous system.

DNA damage response protein TOPBP1 regulates X chromosome silencing in the mammalian germ line.

Meiotic synapsis and recombination between homologs permits the formation of cross-overs that are essential for generating chromosomally balanced sperm and eggs. In mammals, surveillance mechanisms eliminate meiotic cells with defective synapsis, thereby minimizing transmission of aneuploidy. One such surveillance mechanism is meiotic silencing, the inactivation of genes located on asynapsed chromosomes, via ATR-dependent serine-139 phosphorylation of histone H2AFX (γH2AFX). Stimulation of ATR activity requires direct interaction with an ATR activation domain (AAD)-containing partner. However, which partner facilitates the meiotic silencing properties of ATR is unknown. Focusing on the best-characterized example of meiotic silencing, meiotic sex chromosome inactivation, we reveal this AAD-containing partner to be the DNA damage and checkpoint protein TOPBP1. Conditional TOPBP1 deletion during pachynema causes germ cell elimination associated with defective X chromosome gene silencing and sex chromosome condensation. TOPBP1 is essential for localization to the X chromosome of silencing "sensors," including BRCA1, and effectors, including ATR, γH2AFX, and canonical repressive histone marks. We present evidence that persistent DNA double-strand breaks act as silencing initiation sites. Our study identifies TOPBP1 as a critical factor in meiotic sex chromosome silencing.

Tumor-associated APE1 variant exhibits reduced complementation efficiency but does not promote cancer cell phenotypes.

Base excision repair (BER) is the major pathway for coping with most forms of endogenous DNA damage, and defects in the process have been associated with carcinogenesis. Apurinic/apyrimidinic endonuclease 1 (APE1) is a central participant in BER, functioning as a critical endonuclease in the processing of noncoding abasic sites in DNA. Evidence has suggested that APE1 missense mutants, as well as altered expression or localization of the protein, can contribute to disease manifestation. We report herein that the tumor-associated APE1 variant, R237C, shows reduced complementation efficiency of the methyl methanesulfonate hypersensitivity and impaired cell growth exhibited by APE1-deficient mouse embryonic fibroblasts. Overexpression of wild-type APE1 or the R237C variant in the nontransformed C127I mouse cell line had no effect on proliferation, cell cycle status, steady-state DNA damage levels, mitochondrial function, or cellular transformation. A human cell line heterozygous for an APE1 knockout allele had lower levels of endogenous APE1, increased cellular sensitivity to DNA-damaging agents, impaired proliferation with time, and a distinct global gene expression pattern consistent with a stress phenotype. Our results indicate that: (i) the tumor-associated R237C variant is a possible susceptibility factor, but not likely a driver of cancer cell phenotypes, (ii) overexpression of APE1 does not readily promote cellular transformation, and (iii) haploinsufficiency at the APE1 locus can have profound cellular consequences, consistent with BER playing a critical role in proliferating cells. Environ. Mol. Mutagen. 58:84-98, 2017. © 2017 Wiley Periodicals, Inc.

DNA-PKcs, ATM, and ATR Interplay Maintains Genome Integrity during Neurogenesis.

The DNA damage response (DDR) orchestrates a network of cellular processes that integrates cell-cycle control and DNA repair or apoptosis, which serves to maintain genome stability. DNA-PKcs (the catalytic subunit of the DNA-dependent kinase, encoded by PRKDC), ATM (ataxia telangiectasia, mutated), and ATR (ATM and Rad3-related) are related PI3K-like protein kinases and central regulators of the DDR. Defects in these kinases have been linked to neurodegenerative or neurodevelopmental syndromes. In all cases, the key neuroprotective function of these kinases is uncertain. It also remains unclear how interactions between the three DNA damage-responsive kinases coordinate genome stability, particularly in a physiological context. Here, we used a genetic approach to identify the neural function of DNA-PKcs and the interplay between ATM and ATR during neurogenesis. We found that DNA-PKcs loss in the mouse sensitized neuronal progenitors to apoptosis after ionizing radiation because of excessive DNA damage. DNA-PKcs was also required to prevent endogenous DNA damage accumulation throughout the adult brain. In contrast, ATR coordinated the DDR during neurogenesis to direct apoptosis in cycling neural progenitors, whereas ATM regulated apoptosis in both proliferative and noncycling cells. We also found that ATR controls a DNA damage-induced G2/M checkpoint in cortical progenitors, independent of ATM and DNA-PKcs. These nonoverlapping roles were further confirmed via sustained murine embryonic or cortical development after all three kinases were simultaneously inactivated. Thus, our results illustrate how DNA-PKcs, ATM, and ATR have unique and essential roles during the DDR, collectively ensuring comprehensive genome maintenance in the nervous system. SIGNIFICANCE STATEMENT: The DNA damage response (DDR) is essential for prevention of a broad spectrum of different human neurologic diseases. However, a detailed understanding of the DDR at a physiological level is lacking. In contrast to many in vitro cellular studies, here we demonstrate independent biological roles for the DDR kinases DNA-PKcs, ATM, and ATR during neurogenesis. We show that DNA-PKcs is central to DNA repair in nonproliferating cells, and restricts DNA damage accumulation, whereas ATR controls damage-induced G2 checkpoint control and apoptosis in proliferating cells. Conversely, ATM is critical for controlling apoptosis in immature noncycling neural cells after DNA damage. These data demonstrate functionally distinct, but cooperative, roles for each kinase in preserving genome stability in the nervous system.

Cold, clumpy accretion onto an active supermassive black hole.

Supermassive black holes in galaxy centres can grow by the accretion of gas, liberating energy that might regulate star formation on galaxy-wide scales. The nature of the gaseous fuel reservoirs that power black hole growth is nevertheless largely unconstrained by observations, and is instead routinely simplified as a smooth, spherical inflow of very hot gas. Recent theory and simulations instead predict that accretion can be dominated by a stochastic, clumpy distribution of very cold molecular clouds--a departure from the 'hot mode' accretion model--although unambiguous observational support for this prediction remains elusive. Here we report observations that reveal a cold, clumpy accretion flow towards a supermassive black hole fuel reservoir in the nucleus of the Abell 2597 Brightest Cluster Galaxy (BCG), a nearby (redshift z = 0.0821) giant elliptical galaxy surrounded by a dense halo of hot plasma. Under the right conditions, thermal instabilities produce a rain of cold clouds that fall towards the galaxy's centre, sustaining star formation amid a kiloparsec-scale molecular nebula that is found at its core. The observations show that these cold clouds also fuel black hole accretion, revealing 'shadows' cast by the molecular clouds as they move inward at about 300 kilometres per second towards the active supermassive black hole, which serves as a bright backlight. Corroborating evidence from prior observations of warmer atomic gas at extremely high spatial resolution, along with simple arguments based on geometry and probability, indicate that these clouds are within the innermost hundred parsecs of the black hole, and falling closer towards it.

Polynucleotide kinase-phosphatase enables neurogenesis via multiple DNA repair pathways to maintain genome stability.

Polynucleotide kinase-phosphatase (PNKP) is a DNA repair factor possessing both 5'-kinase and 3'-phosphatase activities to modify ends of a DNA break prior to ligation. Recently, decreased PNKP levels were identified as the cause of severe neuropathology present in the human microcephaly with seizures (MCSZ) syndrome. Utilizing novel murine Pnkp alleles that attenuate expression and a T424GfsX48 frame-shift allele identified in MCSZ individuals, we determined how PNKP inactivation impacts neurogenesis. Mice with PNKP inactivation in neural progenitors manifest neurodevelopmental abnormalities and postnatal death. This severe phenotype involved defective base excision repair and non-homologous end-joining, pathways required for repair of both DNA single- and double-strand breaks. Although mice homozygous for the T424GfsX48 allele were lethal embryonically, attenuated PNKP levels (akin to MCSZ) showed general neurodevelopmental defects, including microcephaly, indicating a critical developmental PNKP threshold. Directed postnatal neural inactivation of PNKP affected specific subpopulations including oligodendrocytes, indicating a broad requirement for genome maintenance, both during and after neurogenesis. These data illuminate the basis for selective neural vulnerability in DNA repair deficiency disease.

Aberrant topoisomerase-1 DNA lesions are pathogenic in neurodegenerative genome instability syndromes.

DNA damage is considered to be a prime factor in several spinocerebellar neurodegenerative diseases; however, the DNA lesions underpinning disease etiology are unknown. We observed the endogenous accumulation of pathogenic topoisomerase-1 (Top1)-DNA cleavage complexes (Top1ccs) in murine models of ataxia telangiectasia and spinocerebellar ataxia with axonal neuropathy 1. We found that the defective DNA damage response factors in these two diseases cooperatively modulated Top1cc turnover in a non-epistatic and ATM kinase-independent manner. Furthermore, coincident neural inactivation of ATM and DNA single-strand break repair factors, including tyrosyl-DNA phosphodiesterase-1 or XRCC1, resulted in increased Top1cc formation and excessive DNA damage and neurodevelopmental defects. Notably, direct Top1 poisoning to elevate Top1cc levels phenocopied the neuropathology of the mouse models described above. Our results identify a critical endogenous pathogenic lesion associated with neurodegenerative syndromes arising from DNA repair deficiency, indicating that genome integrity is important for preventing disease in the nervous system.

Neurogenesis requires TopBP1 to prevent catastrophic replicative DNA damage in early progenitors.

The rapid proliferation of progenitors during neurogenesis requires a stringent genomic maintenance program to ensure transmission of genetic fidelity. However the essential factors that govern neural progenitor genome integrity are unknown. Here we report that conditional inactivation of mouse TopBP1, a protein linked to DNA replication, and a key activator of the DNA damage response kinase ATR (ataxia telangiectasia and rad3-related) is critical for maintenance of early-born neural progenitors. During cortical development TopBP1 prevented replication-associated DNA damage in Emx1-progenitors which otherwise resulted in profound tissue ablation. Notably, disrupted neurogenesis in TopBP1-depleted tissues was substantially rescued by inactivation of p53 but not of ATM. Our data establish that TopBP1 is essential for preventing replication-associated DNA strand breaks, but is not essential per se for DNA replication. Thus, TopBP1 is crucial for maintaining genome integrity in the early progenitors that drive neurogenesis.

ATR maintains select progenitors during nervous system development.

The ATR (ATM (ataxia telangiectasia mutated) and rad3-related) checkpoint kinase is considered critical for signalling DNA replication stress and its dysfunction can lead to the neurodevelopmental disorder, ATR-Seckel syndrome. To understand how ATR functions during neurogenesis, we conditionally deleted Atr broadly throughout the murine nervous system, or in a restricted manner in the dorsal telencephalon. Unexpectedly, in both scenarios, Atr loss impacted neurogenesis relatively late during neural development involving only certain progenitor populations. Whereas the Atr-deficient embryonic cerebellar external germinal layer underwent p53- (and p16(Ink4a/Arf))-independent proliferation arrest, other brain regions suffered apoptosis that was partially p53 dependent. In contrast to other organs, in the nervous system, p53 loss did not worsen the outcome of Atr inactivation. Coincident inactivation of Atm also did not affect the phenotype after Atr deletion, supporting non-overlapping physiological roles for these related DNA damage-response kinases in the brain. Rather than an essential general role in preventing replication stress, our data indicate that ATR functions to monitor genomic integrity in a selective spatiotemporal manner during neurogenesis.

DNA ligase III is critical for mtDNA integrity but not Xrcc1-mediated nuclear DNA repair.

DNA replication and repair in mammalian cells involves three distinct DNA ligases: ligase I (Lig1), ligase III (Lig3) and ligase IV (Lig4). Lig3 is considered a key ligase during base excision repair because its stability depends upon its nuclear binding partner Xrcc1, a critical factor for this DNA repair pathway. Lig3 is also present in the mitochondria, where its role in mitochondrial DNA (mtDNA) maintenance is independent of Xrcc1 (ref. 4). However, the biological role of Lig3 is unclear as inactivation of murine Lig3 results in early embryonic lethality. Here we report that Lig3 is essential for mtDNA integrity but dispensable for nuclear DNA repair. Inactivation of Lig3 in the mouse nervous system resulted in mtDNA loss leading to profound mitochondrial dysfunction, disruption of cellular homeostasis and incapacitating ataxia. Similarly, inactivation of Lig3 in cardiac muscle resulted in mitochondrial dysfunction and defective heart-pump function leading to heart failure. However, Lig3 inactivation did not result in nuclear DNA repair deficiency, indicating essential DNA repair functions of Xrcc1 can occur in the absence of Lig3. Instead, we found that Lig1 was critical for DNA repair, but acted in a cooperative manner with Lig3. Additionally, Lig3 deficiency did not recapitulate the hallmark features of neural Xrcc1 inactivation such as DNA damage-induced cerebellar interneuron loss, further underscoring functional separation of these DNA repair factors. Therefore, our data reveal that the critical biological role of Lig3 is to maintain mtDNA integrity and not Xrcc1-dependent DNA repair.

The genesis of cerebellar interneurons and the prevention of neural DNA damage require XRCC1.

Defective responses to DNA single strand breaks underlie various neurodegenerative diseases. However, the exact role of this repair pathway during the development and maintenance of the nervous system is unclear. Using murine neural-specific inactivation of Xrcc1, a factor that is critical for the repair of DNA single strand breaks, we found a profound neuropathology that is characterized by the loss of cerebellar interneurons. This cell loss was linked to p53-dependent cell cycle arrest and occurred as interneuron progenitors commenced differentiation. Loss of Xrcc1 also led to the persistence of DNA strand breaks throughout the nervous system and abnormal hippocampal function. Collectively, these data detail the in vivo link between DNA single strand break repair and neurogenesis and highlight the diverse consequences of specific types of genotoxic stress in the nervous system.

Recurrent genomic alterations characterize medulloblastoma arising from DNA double-strand break repair deficiency.

Inactivation of homologous recombination (HR) or nonhomologous end-joining (NHEJ) predisposes to a spectrum of tumor types. Here, we inactivated DNA double-strand break repair (DSBR) proteins, DNA Ligase IV (Lig4), Xrcc2, and Brca2, or combined Lig4/Xrcc2 during neural development using Nestin-cre. In all cases, inactivation of these repair factors, together with p53 loss, led to rapid medulloblastoma formation. Genomic analysis of these tumors showed recurring chromosome 13 alterations via chromosomal loss or translocations involving regions containing Ptch1. Sequence analysis of the remaining Ptch1 allele showed a variety of inactivating mutations in all tumors analyzed, highlighting the critical tumor suppressor function of this hedgehog-signaling regulator. We also observed genomic amplification or up-regulation of either N-Myc or cyclin D2 in all medulloblastomas. Additionally, chromosome 19, which contains Pten, was also selectively deleted in medulloblastoma arising after disruption of HR. Thus, our data highlight the preeminence of Ptch1 as a tumor suppressor in cerebellar granule cells and reveal other genomic events central to the genesis of medulloblastoma.

Differential DNA damage signaling accounts for distinct neural apoptotic responses in ATLD and NBS.

The MRN complex (Mre11/RAD50/NBS1) and ATM (ataxia telangiectasia, mutated) are critical for the cellular response to DNA damage. ATM disruption causes ataxia telangiectasia (A-T), while MRN dysfunction can lead to A-T-like disease (ATLD) or Nijmegen breakage syndrome (NBS). Neuropathology is a hallmark of these diseases, whereby neurodegeneration occurs in A-T and ATLD while microcephaly characterizes NBS. To understand the contrasting neuropathology resulting from Mre11 or Nbs1 hypomorphic mutations, we analyzed neural tissue from Mre11(ATLD1/ATLD1) and Nbs1(DeltaB/DeltaB) mice after genotoxic stress. We found a pronounced resistance to DNA damage-induced apoptosis after ionizing radiation or DNA ligase IV (Lig4) loss in the Mre11(ATLD1/ATLD1) nervous system that was associated with defective Atm activation and phosphorylation of its substrates Chk2 and p53. Conversely, DNA damage-induced Atm phosphorylation was defective in Nbs1(DeltaB/DeltaB) neural tissue, although apoptosis occurred normally. We also conditionally disrupted Lig4 throughout the nervous system using Nestin-cre (Lig4(Nes-Cre)), and while viable, these mice showed pronounced microcephaly and a prominent age-related accumulation of DNA damage throughout the brain. Either Atm-/- or Mre11(ATLD1/ATLD1) genetic backgrounds, but not Nbs1(DeltaB/DeltaB), rescued Lig4(Nes-Cre) microcephaly. Thus, DNA damage signaling in the nervous system is different between ATLD and NBS and likely explains their respective neuropathology.

Scythe regulates apoptosis-inducing factor stability during endoplasmic reticulum stress-induced apoptosis.

Scythe (BAT3; HLA-B associated transcript 3, Bag 6) is a protein that has been implicated in apoptosis because it can modulate the Drosophila melanogaster apoptotic regulator, Reaper. Mice lacking Scythe show pronounced defects in organogenesis and in the regulation of apoptosis and proliferation during mammalian development. However, the biochemical pathways important for Scythe function are unknown. We report here multiple levels of interaction between Scythe and the apoptogenic mitochondrial intermembrane protein AIF (apoptosis-inducing factor). Scythe physically interacts with AIF and regulates its stability. AIF stability is markedly reduced in Scythe(-/-) cells, which are more resistant to endoplasmic reticulum stress induced by thapsigargin. Reintroduction of Scythe or overexpression of AIF in Scythe(-/-) cells restores their sensitivity to apoptosis. Together, these data implicate Scythe as a regulator of AIF.

TDP1 facilitates chromosomal single-strand break repair in neurons and is neuroprotective in vivo.

Defective Tyrosyl-DNA phosphodiesterase 1 (TDP1) can cause spinocerebellar ataxia with axonal neuropathy (SCAN1), a neurodegenerative syndrome associated with marked cerebellar atrophy and peripheral neuropathy. Although SCAN1 lymphoblastoid cells show pronounced defects in the repair of chromosomal single-strand breaks (SSBs), it is unknown if this DNA repair activity is important for neurons or for preventing neurodegeneration. Therefore, we generated Tdp1-/- mice to assess the role of Tdp1 in the nervous system. Using both in vitro and in vivo assays, we found that cerebellar neurons or primary astrocytes derived from Tdp1-/- mice display an inability to rapidly repair DNA SSBs associated with Top1-DNA complexes or oxidative damage. Moreover, loss of Tdp1 resulted in age-dependent and progressive cerebellar atrophy. Tdp1-/- mice treated with topotecan, a drug that increases levels of Top1-DNA complexes, also demonstrated significant loss of intestinal and hematopoietic progenitor cells. These data indicate that TDP1 is required for neural homeostasis, and reveal a widespread requisite for TDP1 function in response to acutely elevated levels of Top1-associated DNA strand breaks.

Patched2 modulates tumorigenesis in patched1 heterozygous mice.

The sonic hedgehog (SHH) receptor Patched 1 (Ptch1) is critical for embryonic development, and its loss is linked to tumorigenesis. Germ line inactivation of one copy of Ptch1 predisposes to basal cell carcinoma and medulloblastoma in mouse and man. In many cases, medulloblastoma arising from perturbations of Ptch1 function leads to a concomitant up-regulation of a highly similar gene, Patched2 (Ptch2). As increased expression of Ptch2 is associated with medulloblastoma and other tumors, we investigated the role of Ptch2 in tumor suppression by generating Ptch2-deficient mice. In striking contrast to Ptch1-/- mice, Ptch2-/- animals were born alive and showed no obvious defects and were not cancer prone. However, loss of Ptch2 markedly affected tumor formation in combination with Ptch1 haploinsufficiency. Ptch1+/-Ptch2-/- and Ptch1+/-Ptch2+/- animals showed a higher incidence of tumors and a broader spectrum of tumor types compared with Ptch1+/- animals. Therefore, Ptch2 modulates tumorigenesis associated with Ptch1 haploinsufficiency.