Agrin-Lrp4-Ror2 signaling regulates adult hippocampal neurogenesis in mice.

Adult neurogenesis in the hippocampus may represent a form of plasticity in brain functions including mood, learning and memory. However, mechanisms underlying neural stem/progenitor cells (NSPCs) proliferation are not well understood. We found that Agrin, a factor critical for neuromuscular junction formation, is elevated in the hippocampus of mice that are stimulated by enriched environment (EE). Genetic deletion of the Agrn gene in excitatory neurons decreases NSPCs proliferation and increases depressive-like behavior. Low-density lipoprotein receptor-related protein 4 (Lrp4), a receptor for Agrin, is expressed in hippocampal NSPCs and its mutation blocked basal as well as EE-induced NSPCs proliferation and maturation of newborn neurons. Finally, we show that Lrp4 interacts with and activates receptor tyrosine kinase-like orphan receptor 2 (Ror2); and Ror2 mutation impairs NSPCs proliferation. Together, these observations identify a role of Agrin-Lrp4-Ror2 signaling for adult neurogenesis, uncovering previously unexpected functions of Agrin and Lrp4 in the brain.

A Wnt/Calcium Signaling Cascade Regulates Neuronal Excitability and Trafficking of NMDARs.

Wnt signaling controls multiple biological process, particularly the embryonic development of metazoans. Sustained expression of Wnt signaling components in the mature mammalian CNS and their apparent deregulation in certain neuropathologies suggest that it also plays a part beyond embryonic development to regulate normal brain function. We describe a noncanonical Wnt/Ca2+ signaling cascade that regulates the electrophysiological intrinsic properties of rat neurons, resulting in sustained membrane depolarization and the mobilization of Ca2+ from internal stores. These effects require tyrosine kinase-like orphan receptor 2 (RoR2), activation of PLC, and voltage-gated Ca2+ channels. Activation of this signaling cascade then promotes surface expression of N-methyl-D-aspartate receptors (NMDARs) through a SNARE-dependent mechanism. This neuronal Wnt/Ca2+ signaling pathway represents a mechanism for Wnt ligands to regulate normal brain processes in the mature animal and provides a framework for understanding how alterations in this pathway may contribute to the etiology of psychiatric disorders where NMDARs are compromised.

Planar cell polarity signaling in the uterus directs appropriate positioning of the crypt for embryo implantation.

Blastocyst implantation is a complex process requiring coordination of a dynamic sequence of embryo-uterine interactions. Blood vessels enter the uterus from the mesometrium, demarcating the uterus into mesometrial (M) and antimesometrial (AM) domains. Implantation occurs along the uterine longitudinal axis within specialized implantation chambers (crypts) that originate within the evaginations directed from the primary lumen toward the AM domain. The morphological orientation of crypts in rodent uteri was recognized more than a century ago, but the mechanism remained unknown. Here we provide evidence that planar cell polarity (PCP) signaling orchestrates directed epithelial evaginations to form crypts for implantation in mice. Uterine deletion of Vang-like protein 2, but not Vang-like protein 1, conferred aberrant PCP signaling, misdirected epithelial evaginations, defective crypt formation, and blastocyst attachment, leading to severely compromised pregnancy outcomes. The study reveals a previously unrecognized role for PCP in executing spatial cues for crypt formation and implantation. Because PCP is an evolutionarily conserved phenomenon, our study is likely to inspire implantation studies of this signaling pathway in humans and other species.

Targeting ROR1 inhibits the self-renewal and invasive ability of glioblastoma stem cells.

Glioblastoma is the most malignant of brain tumours and is difficult to cure because of interruption of drug delivery by the blood-brain barrier system, its high metastatic capacity and the existence of cancer stem cells (CSCs). Although CSCs are present as a small population in malignant tumours, CSCs have been studied as they are responsible for causing recurrence, metastasis and resistance to chemotherapy and radiotherapy for cancer. CSCs have self-renewal characteristics like normal stem cells. The aim of this study was to investigate whether receptor tyrosine kinase-like orphan receptor 1 (ROR1) is involved in stem cell maintenance and malignant properties in human glioblastoma. Knockdown of ROR1 caused reduction of stemness and sphere formation capacity. Moreover, down-regulation of ROR1 suppressed the expression of epithelial-mesenchymal transition-related genes and the tumour migratory and invasive abilities. The results of this study indicate that targeting ROR1 can induce differentiation of CSCs and inhibit metastasis in glioblastoma. In addition, ROR1 may be used as a potential marker for glioblastoma stem cells as well as a potential target for glioblastoma stem cell therapy.

Inhibition of the receptor tyrosine kinase ROR1 by anti-ROR1 monoclonal antibodies and siRNA induced apoptosis of melanoma cells.

The receptor tyrosine kinase (RTK) ROR1 is overexpressed and of importance for the survival of various malignancies, including lung adenocarcinoma, breast cancer and chronic lymphocytic leukemia (CLL). There is limited information however on ROR1 in melanoma. In the present study we analysed in seven melanoma cell lines ROR1 expression and phosphorylation as well as the effects of anti-ROR1 monoclonal antibodies (mAbs) and ROR1 suppressing siRNA on cell survival. ROR1 was overexpressed at the protein level to a varying degree and phosphorylated at tyrosine and serine residues. Three of our four self-produced anti-ROR1 mAbs (clones 3H9, 5F1 and 1A8) induced a significant direct apoptosis of the ESTDAB049, ESTDAB112, DFW and A375 cell lines as well as cell death in complement dependent cytotoxicity (CDC) and antibody dependent cellular cytotoxicity (ADCC). The ESTDAB081 and 094 cell lines respectively were resistant to direct apoptosis of the four anti-ROR1 mAbs alone but not in CDC or ADCC. ROR1 siRNA transfection induced downregulation of ROR1 expression both at mRNA and protein levels proceeded by apoptosis of the melanoma cells (ESTDAB049, ESTDAB112, DFW and A375) including ESTDAB081, which was resistant to the direct apoptotic effect of the mAbs. The results indicate that ROR1 may play a role in the survival of melanoma cells. The surface expression of ROR1 on melanoma cells may support the notion that ROR1 might be a suitable target for mAb therapy.

Wnt5a-Ror2 signaling between osteoblast-lineage cells and osteoclast precursors enhances osteoclastogenesis.

The signaling molecule Wnt regulates bone homeostasis through ß-catenin-dependent canonical and ß-catenin-independent noncanonical pathways. Impairment of canonical Wnt signaling causes bone loss in arthritis and osteoporosis; however, it is unclear how noncanonical Wnt signaling regulates bone resorption. Wnt5a activates noncanonical Wnt signaling through receptor tyrosine kinase-like orphan receptor (Ror) proteins. We showed that Wnt5a-Ror2 signaling between osteoblast-lineage cells and osteoclast precursors enhanced osteoclastogenesis. Osteoblast-lineage cells expressed Wnt5a, whereas osteoclast precursors expressed Ror2. Mice deficient in either Wnt5a or Ror2, and those with either osteoclast precursor-specific Ror2 deficiency or osteoblast-lineage cell-specific Wnt5a deficiency showed impaired osteoclastogenesis. Wnt5a-Ror2 signals enhanced receptor activator of nuclear factor-κB (RANK) expression in osteoclast precursors by activating JNK and recruiting c-Jun on the promoter of the gene encoding RANK, thereby enhancing RANK ligand (RANKL)-induced osteoclastogenesis. A soluble form of Ror2 acted as a decoy receptor of Wnt5a and abrogated bone destruction in mouse arthritis models. Our results suggest that the Wnt5a-Ror2 pathway is crucial for osteoclastogenesis in physiological and pathological environments and represents a therapeutic target for bone diseases, including arthritis.

Mice lacking the orphan receptor ror1 have distinct skeletal abnormalities and are growth retarded.

Ror1 is a member of the Ror-family receptor tyrosine kinases. Ror1 is broadly expressed in various tissues and organs during mouse embryonic development. However, so far little is known about its function. The closely related family member Ror2 was shown to play a crucial role in skeletogenesis and has been shown to act as a co-receptor for Wnt5a mediating non-canonical Wnt-signaling. Previously, it has been shown that during embryonic development Ror1 acts in part redundantly with Ror2 in the skeletal and cardiovascular systems. In this study, we report that loss of the orphan receptor Ror1 results in a variety of phenotypic defects within the skeletal and urogenital systems and that Ror1 mutant mice display a postnatal growth retardation phenotype.

Ror2 receptor requires tyrosine kinase activity to mediate Wnt5A signaling.

The Wnts include a large family of secreted proteins that serve as important signals during embryonic development and adult homeostasis. In the most well understood Wnt signaling pathway, Wnt binding to Frizzled and low density lipoprotein receptor-related protein induces beta-catenin protein stabilization and entry into the nucleus, resulting in changes in target gene transcription. Emerging evidence suggests that Wnt5a can inhibit Wnt/beta-catenin signaling through interaction with the receptor Ror2. The Ror2 protein belongs to the receptor tyrosine kinase superfamily and contains several recognizable structural motifs. However, limited information is available regarding which specific domains are required for the inhibitory signaling activity of Wnt5a. Through mutation and deletion analysis, we have analyzed which specific domains and residues, including those necessary for tyrosine kinase activity, mediate the Wnt5a signal. To determine whether Ror2 can inhibit canonical Wnt signaling in vivo, we examined the effect of Ror2 loss on the expression of the Wnt reporter Axin2(LacZ), finding increased reporter activity in Ror2 null mice, demonstrating that Ror2 can also inhibit Wnt/beta-catenin signaling in the context of intact tissues.