The USP7-TRIM27 axis mediates non-canonical PRC1.1 function and is a druggable target in leukemia

In an attempt to unravel functionality of the non-canonical PRC1.1 Polycomb complex in human leukemogenesis, we show that USP7 and TRIM27 are integral components of PRC1.1. USP7 interactome analyses show that PRC1.1 is the predominant Polycomb complex co-precipitating with USP7. USP7 inhibition results in PRC1.1 disassembly and loss of chromatin binding, coinciding with reduced H2AK119ub and H3K27ac levels and diminished gene transcription of active PRC1.1-controlled loci, whereas H2AK119ub marks are also lost at PRC1 loci. TRIM27 and USP7 are reciprocally required for incorporation into PRC1.1, and TRIM27 knockdown partially rescues USP7 inhibitor sensitivity. USP7 inhibitors effectively impair proliferation in AML cells in vitro, also independent of the USP7-MDM2-TP53 axis, and MLL-AF9-induced leukemia is delayed in vivo in human leukemia xenografts. We propose a model where USP7 counteracts TRIM27 E3 ligase activity, thereby maintaining PRC1.1 integrity and function. Moreover, USP7 inhibition may be a promising new strategy to treat AML patients

LKB1 Destabilizes Microtubules in Myoblasts and Contributes to Myoblast Differentiation

Background: Skeletal muscle myoblast differentiation and fusion into multinucleate myotubes is associated with dramatic cytoskeletal changes. We find that microtubules in differentiated myotubes are highly stabilized, but premature microtubule stabilization blocks differentiation. Factors responsible for microtubule destabilization in myoblasts have not been identified. Findings: We find that a transient decrease in microtubule stabilization early during myoblast differentiation precedes the ultimate microtubule stabilization seen in differentiated myotubes. We report a role for the serine-threonine kinase LKB1 in both microtubule destabilization and myoblast differentiation. LKB1 overexpression reduced microtubule elongation in a Nocodazole washout assay, and LKB1 RNAi increased it, showing LKB1 destabilizes microtubule assembly in myoblasts. LKB1 levels and activity increased during myoblast differentiation, along with activation of the known LKB1 substrates AMPactivated protein kinase (AMPK) and microtubule affinity regulating kinases (MARKs). LKB1 overexpression accelerated differentiation, whereas RNAi impaired it. Conclusions: Reduced microtubule stability precedes myoblast differentiation and the associated ultimate microtubule stabilization seen in myotubes. LKB1 plays a positive role in microtubule destabilization in myoblasts and in myoblast differentiation. This work suggests a model by which LKB1-induced microtubule destabilization facilitates the cytoskeleta

CLP1 Founder Mutation Links tRNA Splicing and Maturation to Cerebellar Development and Neurodegeneration

SummaryNeurodegenerative diseases can occur so early as to affect neurodevelopment. From a cohort of more than 2,000 consanguineous families with childhood neurological disease, we identified a founder mutation in four independent pedigrees in cleavage and polyadenylation factor I subunit 1 (CLP1). CLP1 is a multifunctional kinase implicated in tRNA, mRNA, and siRNA maturation. Kinase activity of the CLP1 mutant protein was defective, and the tRNA endonuclease complex (TSEN) was destabilized, resulting in impaired pre-tRNA cleavage. Germline clp1 null zebrafish showed cerebellar neurodegeneration that was rescued by wild-type, but not mutant, human CLP1 expression. Patient-derived induced neurons displayed both depletion of mature tRNAs and accumulation of unspliced pre-tRNAs. Transfection of partially processed tRNA fragments into patient cells exacerbated an oxidative stress-induced reduction in cell survival. Our data link tRNA maturation to neuronal development and neurodegeneration through defective CLP1 function in humans

LKB1/AMPK and PKA Control ABCB11 Trafficking and Polarization in Hepatocytes.

Polarization of hepatocytes is manifested by bile canalicular network formation and activation of LKB1 and AMPK, which control cellular energy metabolism. The bile acid, taurocholate, also regulates development of the canalicular network through activation of AMPK. In the present study, we used collagen sandwich hepatocyte cultures from control and liver-specific LKB1 knockout mice to examine the role of LKB1 in trafficking of ABCB11, the canalicular bile acid transporter. In polarized hepatocytes, ABCB11 traffics from Golgi to the apical plasma membrane and endogenously cycles through the rab 11a-myosin Vb recycling endosomal system. LKB1 knockout mice were jaundiced, lost weight and manifested impaired bile canalicular formation and intracellular trafficking of ABCB11, and died within three weeks. Using live cell imaging, fluorescence recovery after photobleaching (FRAP), particle tracking, and biochemistry, we found that LKB1 activity is required for microtubule-dependent trafficking of ABCB11 to the canalicular membrane. In control hepatocytes, ABCB11 trafficking was accelerated by taurocholate and cAMP; however, in LKB1 knockout hepatocytes, ABCB11 trafficking to the apical membrane was greatly reduced and restored only by cAMP, but not taurocholate. cAMP acted through a PKA-mediated pathway which did not activate AMPK. Our studies establish a regulatory role for LKB1 in ABCB11 trafficking to the canalicular membrane, hepatocyte polarization, and canalicular network formation

A review of the dodo and its ecosystem: insights from a vertebrate concentration Lagerstätte in Mauritius

The dodo Raphus cucullatus Linnaeus, an extinct and flightless, giant pigeon endemic to Mauritius, has fascinated people since its discovery, yet has remained surprisingly poorly known. Until the mid-19th century, almost all that was known about the dodo was based on illustrations and written accounts by 17th century mariners, often of questionable accuracy. Furthermore, only a few fragmentary remains of dodos collected prior to the bird's extinction exist. Our understanding of the dodo's anatomy was substantially enhanced by the discovery in 1865 of subfossil bones in a marsh called the Mare aux Songes, situated in southeastern Mauritius. However, no contextual information was recorded during early excavation efforts, and the majority of excavated material comprised larger dodo bones, almost all of which were unassociated. Here we present a modern interdisciplinary analysis of the Mare aux Songes, a 4200-year-old multitaxic vertebrate concentration Lagerstätte. Our analysis of the deposits at this site provides the first detailed overview of the ecosystem inhabited by the dodo. The interplay of climatic and geological conditions led to the exceptional preservation of the animal and associated plant remains at the Mare aux Songes and provides a window into the past ecosystem of Mauritius. This interdisciplinary research approach provides an ecological framework for the dodo, complementing insights on its anatomy derived from the only associated dodo skeletons known, both of which were collected by Etienne Thirioux and are the primary subject of this memoir.publishedVersio

Congenital Hydrocephalus and Abnormal Subcommissural Organ Development in Sox3 Transgenic Mice

Congenital hydrocephalus (CH) is a life-threatening medical condition in which excessive accumulation of CSF leads to ventricular expansion and increased intracranial pressure. Stenosis (blockage) of the Sylvian aqueduct (Aq; the narrow passageway that connects the third and fourth ventricles) is a common form of CH in humans, although the genetic basis of this condition is unknown. Mouse models of CH indicate that Aq stenosis is associated with abnormal development of the subcommmissural organ (SCO) a small secretory organ located at the dorsal midline of the caudal diencephalon. Glycoproteins secreted by the SCO generate Reissner's fibre (RF), a thread-like structure that descends into the Aq and is thought to maintain its patency. However, despite the importance of SCO function in CSF homeostasis, the genetic program that controls SCO development is poorly understood. Here, we show that the X-linked transcription factor SOX3 is expressed in the murine SCO throughout its development and in the mature organ. Importantly, overexpression of Sox3 in the dorsal diencephalic midline of transgenic mice induces CH via a dose-dependent mechanism. Histological, gene expression and cellular proliferation studies indicate that Sox3 overexpression disrupts the development of the SCO primordium through inhibition of diencephalic roof plate identity without inducing programmed cell death. This study provides further evidence that SCO function is essential for the prevention of hydrocephalus and indicates that overexpression of Sox3 in the dorsal midline alters progenitor cell differentiation in a dose-dependent manner

NOS1AP is a novel molecular target and critical factor in TDP-43 pathology

Cappelli et al. reported that Nitric Oxide Synthase 1 Adaptor Protein is a co-regulated transcript of the TAR DNA-binding protein 43 kDa, reduced in amyotrophic lateral sclerosis and frontotemporal lobar degeneration patients with TAR DNA-binding protein 43 kDa pathology. Overall, their results highlight Nitric Oxide Synthase 1 Adaptor Protein as a novel druggable disease-relevant gene in TAR DNA-binding protein 43 kDa-related proteinopathies.Many lines of evidence have highlighted the role played by heterogeneous nuclear ribonucleoproteins in amyotrophic lateral sclerosis. In this study, we have aimed to identify transcripts co-regulated by TAR DNA-binding protein 43 kDa and highly conserved heterogeneous nuclear ribonucleoproteins which have been previously shown to regulate TAR DNA-binding protein 43 kDa toxicity (deleted in azoospermia-associated protein 1, heterogeneous nuclear ribonucleoprotein -Q, -D, -K and -U). Using the transcriptome analyses, we have uncovered that Nitric Oxide Synthase 1 Adaptor Protein mRNA is a direct TAR DNA-binding protein 43 kDa target, and in flies, its modulation alone can rescue TAR DNA-binding protein 43 kDa pathology. In primary mouse cortical neurons, we show that TAR DNA-binding protein 43 kDa mediated downregulation of Nitric Oxide Synthase 1 Adaptor Protein expression strongly affects the NMDA-receptor signalling pathway. In human patients, the downregulation of Nitric Oxide Synthase 1 Adaptor Protein mRNA strongly correlates with TAR DNA-binding protein 43 kDa proteinopathy as measured by cryptic Stathmin-2 and Unc-13 homolog A cryptic exon inclusion. Overall, our results demonstrate that Nitric Oxide Synthase 1 Adaptor Protein may represent a novel disease-relevant gene, potentially suitable for the development of new therapeutic strategies

Rare variant analyses validate known ALS genes in a multi-ethnic population and identifies ANTXR2 as a candidate in PLS

BackgroundAmyotrophic lateral sclerosis (ALS) is a neurodegenerative disease affecting over 300,000 people worldwide. It is characterized by the progressive decline of the nervous system that leads to the weakening of muscles which impacts physical function. Approximately, 15% of individuals diagnosed with ALS have a known genetic variant that contributes to their disease. As therapies that slow or prevent symptoms continue to develop, such as antisense oligonucleotides, it is important to discover novel genes that could be targets for treatment. Additionally, as cohorts continue to grow, performing analyses in ALS subtypes, such as primary lateral sclerosis (PLS), becomes possible due to an increase in power. These analyses could highlight novel pathways in disease manifestation.MethodsBuilding on our previous discoveries using rare variant association analyses, we conducted rare variant burden testing on a substantially larger multi-ethnic cohort of 6,970 ALS patients, 166 PLS patients, and 22,524 controls. We used intolerant domain percentiles based on sub-region Residual Variation Intolerance Score (subRVIS) that have been described previously in conjunction with gene based collapsing approaches to conduct burden testing to identify genes that associate with ALS and PLS.ResultsA gene based collapsing model showed significant associations with SOD1, TARDBP, and TBK1 (OR = 19.18, p = 3.67 × 10–39; OR = 4.73, p = 2 × 10–10; OR = 2.3, p = 7.49 × 10–9, respectively). These genes have been previously associated with ALS. Additionally, a significant novel control enriched gene, ALKBH3 (p = 4.88 × 10–7), was protective for ALS in this model. An intolerant domain-based collapsing model showed a significant improvement in identifying regions in TARDBP that associated with ALS (OR = 10.08, p = 3.62 × 10–16). Our PLS protein truncating variant collapsing analysis demonstrated significant case enrichment in ANTXR2 (p = 8.38 × 10–6).ConclusionsIn a large multi-ethnic cohort of 6,970 ALS patients, collapsing analyses validated known ALS genes and identified a novel potentially protective gene, ALKBH3. A first-ever analysis in 166 patients with PLS found a candidate association with loss-of-function mutations in ANTXR2