The Mre11 complex (Mre11, Rad50, and Nbs1) is a central component of the DNA damage response (DDR), governing both double-strand break repair and DDR signaling. Rad50 contains a highly conserved Zn 2+ -dependent homodimerization interface, the Rad50 hook domain. Mutations that inactivate the hook domain produce a null phenotype. In this study, we analyzed mutants with reduced hook domain function in an effort to stratify hookdependent Mre11 complex functions. One of these alleles, Rad50 46 , conferred reduced Zn 2+ affinity and dimerization efficiency. Homozygous Rad50 46/46 mutations were lethal in mice. However, in the presence of wildtype Rad50, Rad50 46 exerted a dominant gain-of-function phenotype associated with chronic DDR signaling. At the organismal level, Rad50 +/46 exhibited hydrocephalus, liver tumorigenesis, and defects in primitive hematopoietic and gametogenic cells. These outcomes were dependent on ATM, as all phenotypes were mitigated in Rad50 +/46 Atm +/-mice. These data reveal that the murine Rad50 hook domain strongly influences Mre11 complex-dependent DDR signaling, tissue homeostasis, and tumorigenesis.

B-cell lymphoma-2 (Bcl-2) family members have been demonstrated to play a crucial role in the regulation of apoptosis as mediators in between the apical stimuli sensing steps and the executory mechanisms of apoptosis. Deregulation of their role may subvert the homeostasis of a given tissue and collaborate in the genesis of a myriad of diseases characterised by exacerbated or insufficient apoptosis, including diseases such as neurodegenerative diseases or cancer. Structural studies have defined homology regions shared by the members of the family that are responsible of the network of interactions established amongst the members of the family. These proteins usually form heterodimers between the so called antiapoptotic multidomain members and the proapoptotic BH3-only proteins. As a consequence, mitochondrial apoptogenic proteins are released to the cytoplasm and the apoptotic signal proceeds towards the final, execution phase of the apoptotic process. The high complexity of the family (more than 20 members have been isolated) makes the study of individual proteins difficult. Genetic approaches have revealed a high degree of redundancy in the family. Only a few proteins belonging to the antiapoptotic group have been proven to be essential for correct embryonic development. Genetic inactivation in mice shows a dramatic phenotype characterised by massive cell death in multiple tissues during embryogenesis, which leads from very early up to perinatal death. This genetic evidence proves the importance of the members of the family for the regulation of apoptosis in order to achieve the proper development and homeostasis of tissues and organs.

Cyclin O (encoded by CCNO) is a member of the cyclin family with regulatory functions in ciliogenesis and apoptosis. Homozygous CCNO mutations have been identified in human patients with Reduced Generation of Multiple Motile Cilia (RGMC) and conditional inactivation of Ccno in the mouse recapitulates some of the pathologies associated with the human disease. These include defects in the development of motile cilia and hydrocephalus. To further investigate the functions of Ccno in vivo, we have generated a new mouse model characterized by the constitutive loss of Ccno in all tissues and followed a cohort during ageing. Ccno-/- mice were growth impaired and developed hydrocephalus with high penetrance. In addition, some Ccno+/- mice also developed hydrocephalus and affected Ccno-/- and Ccno+/- mice exhibited additional CNS defects including cortical thinning and hippocampal abnormalities. In addition to the CNS defects, both male and female Ccno-/- mice were infertile and female mice exhibited few motile cilia in the oviduct. Our results further establish CCNO as an important gene for normal development and suggest that heterozygous CCNO mutations could underlie hydrocephalus or diminished fertility in some human patients.

We have identified an early step common to pathways activated by different forms of intrinsic apoptosis stimuli. It requires de novo synthesis of a novel cyclin, cyclin O, that forms active complexes primarily with Cdk2 upon apoptosis induction in lymphoid cells. Cyclin O expression precedes glucocorticoid and c-radiation-induced apoptosis in vivo in mouse thymus and spleen, and its overexpression induces caspase-dependent apoptosis in cultured cells. Knocking down the endogenous expression of cyclin O by shRNA leads to the inhibition of glucocorticoid and DNA damage-induced apoptosis due to a failure in the activation of apical caspases while leaving CD95 death receptor-mediated apoptosis intact. Our data demonstrate that apoptosis induction in lymphoid cells is one of the physiological roles of cyclin O and it does not act by perturbing a normal cellular process such as the cell cycle, the DNA damage checkpoints or transcriptional response to glucocorticoids.

The MRE11 complex (MRE11, RAD50, and NBS1) is a central component of the DNA damage response, governing both double-strand break repair and DNA damage response signaling. To determine the functions of the MRE11 complex in the development and maintenance of oocytes, we analyzed ovarian phenotypes of mice harboring the hypomorphic Mre11ATLD1 allele. Mre11ATLD1/ATLD1 females exhibited premature oocyte elimination attributable to defects in homologous chromosome pairing and double-strand break repair during meiotic prophase. Other aspects of meiotic progression, including attachment of telomeres to the nuclear envelope and recruitment of RAD21L, a component of the meiotic cohesin complex to the synaptonemal complex, were normal. Unlike Dmc1−/− and Trp13Gt/Gt mice which exhibit comparable defects in double-strand break repair and oocyte depletion by 5 days post-partum, we found that oocyte attrition occurred by 12 weeks in Mre11ATLD1/ATLD1. Disruption of the oocyte checkpoint pathway governed by Chk2 gene further enhanced the survival of Mre11ATLD1/ATLD1 follicles. Together our data suggest that the MRE11 complex influences the elimination of oocytes with unrepaired meiotic double-strand breaks post-natally, in addition to its previously described role in double-strand break repair and homologous synapsis during female meiosis.