Gang Ma

Wls-mediated Wnts differentially regulate distal limb patterning and tissue morphogenesis.

Wnt proteins are diffusible morphogens that play multiple roles during vertebrate limb development. However, the complexity of Wnt signaling cascades and their overlapping expression prevent us from dissecting their function in limb patterning and tissue morphogenesis. Depletion of the Wntless (Wls) gene, which is required for the secretion of various Wnts, makes it possible to genetically dissect the overall effect of Wnts in limb development. In this study, the Wls gene was conditionally depleted in limb mesenchyme and ectoderm. The loss of mesenchymal Wls prevented the differentiation of distal mesenchyme and arrested limb outgrowth, most likely by affecting Wnt5a function. Meanwhile, the deletion of ectodermal Wls resulted in agenesis of distal limb tissue and premature regression of the distal mesenchyme. These observations suggested that Wnts from the two germ layers differentially regulate the pool of undifferentiated distal limb mesenchyme cells. Cellular behavior analysis revealed that ectodermal Wnts sustain mesenchymal cell proliferation and survival in a manner distinct from Fgf. Ectodermal Wnts were also shown for the first time to be essential for distal tendon/ligament induction, myoblast migration and dermis formation in the limb. These findings provide a comprehensive view of the role of Wnts in limb patterning and tissue morphogenesis.

c-Abl promotes osteoblast expansion by differentially regulating canonical and non-canonical BMP pathways and p16INK4a expression

Defects in stem cell renewal or progenitor cell expansion underlie ageing-related diseases such as osteoporosis. Yet much remains unclear about the mechanisms regulating progenitor expansion. Here we show that the tyrosine kinase c-Abl plays an important role in osteoprogenitor expansion. c-Abl interacts with and phosphorylates BMPRIA and the phosphorylation differentially influences the interaction of BMPRIA with BMPRII and the Tab1–Tak1 complex, leading to uneven activation of Smad1/5/8 and Erk1/2, the canonical and non-canonical BMP pathways that direct the expression of p16INK4a. c-Abl deficiency shunts BMP signalling from Smad1/5/8 to Erk1/2, leading to p16INK4a upregulation and osteoblast senescence. Mouse genetic studies revealed that p16INK4a controls mesenchymal stem cell maintenance and osteoblast expansion and mediates the effects of c-Abl deficiency on osteoblast expansion and bone formation. These findings identify c-Abl as a regulator of BMP signalling pathways and uncover a role for c-Abl in p16INK4a expression and osteoprogenitor expansion.

Wnt-mediated reciprocal regulation between cartilage and bone development during endochondral ossification

The role of Wnt signaling is extensively studied in skeletal development and postnatal bone remodeling, mostly based on the genetic approaches of β-catenin manipulation. However, given their independent function, a requirement for β-catenin is not the same as that for Wnt. Here, we investigated the effect of Wnt proteins in both tissues through generating cartilage- or bone-specific Wls null mice, respectively. Depletion of Wls by Col2-Cre, which would block Wnt secretion in the chondrocytes and perichondrium, delayed chondrocyte hypertrophy in the growth plate and impaired perichondrial osteogenesis. Loss of Wls in chondrocytes also disturbed the proliferating chondrocyte morphology and division orientation, which was similar to the defect observed in Wnt5a null mice. On the other hand, inactivation of Wls in osteoblasts by Col1-Cre resulted in a shorter hypertrophic zone and an increase of TRAP positive cell number in the chondro-osseous junction of growth plate, coupled with a decrease in bone mass. Taken together, our studies reveal that Wnt proteins not only modulate differentiation and cellular communication within populations of chondrocytes, but also mediate the cross regulation between the chondrocytes and osteoblasts in growth plate.

Ablation of Tak1 in osteoclast progenitor leads to defects in skeletal growth and bone remodeling in mice

Tak1 is a MAPKKK that can be activated by growth factors and cytokines such as RANKL and BMPs and its downstream pathways include NF-κB and JNK/p38 MAPKs. Tak1 is essential for mouse embryonic development and plays critical roles in tissue homeostasis. Previous studies have shown that Tak1 is a positive regulator of osteoclast maturation, yet its roles in bone growth and remodeling have not been assessed, as mature osteoclast-specific Tak1 deletion with Cstk-Cre resulted in runtedness and postnatal lethality. Here we generated osteoclast progenitor (monocyte)-specific Tak1 knockout mice and found that these mice show normal body weight, limb size and fertility, and osteopetrosis with severity similar to that of RANK or RANKL deficient mice. Mechanistically, Tak1 deficiency altered the signaling of NF-κB, p38MAPK, and Smad1/5/8 and the expression of PU.1, MITF, c-Fos, and NFATc1, suggesting that Tak1 regulates osteoclast differentiation at multiple stages via multiple signaling pathways. Moreover, the Tak1 mutant mice showed defects in skull, articular cartilage, and mesenchymal stromal cells. Ex vivo Tak1−/− monocytes also showed enhanced ability in promoting osteogenic differentiation of mesenchymal stromal cells. These findings indicate that Tak1 functions in osteoclastogenesis in a cell-autonomous manner and in osteoblastogenesis and chondrogenesis in non-cell-autonomous manners.

Ndrg2 regulates vertebral specification in differentiating somites.

It is generally thought that vertebral patterning and identity are globally determined prior to somite formation. Relatively little is known about the regulators of vertebral specification after somite segmentation. Here, we demonstrated that Ndrg2, a tumor suppressor gene, was dynamically expressed in the presomitic mesoderm (PSM) and at early stage of differentiating somites. Loss of Ndrg2 in mice resulted in vertebral homeotic transformations in thoracic/lumbar and lumbar/sacral transitional regions in a dose-dependent manner. Interestingly, the inactivation of Ndrg2 in osteoblasts or chondrocytes caused defects resembling those observed in Ndrg2(-/-) mice, with a lower penetrance. In addition, forced overexpression of Ndrg2 in osteoblasts or chondrocytes also conferred vertebral defects, which were distinct from those in Ndrg2(-/-) mice. These genetic analyses revealed that Ndrg2 modulates vertebral identity in segmented somites rather than in the PSM. At the molecular level, combinatory alterations of the amount of Hoxc8-11 gene transcripts were detected in the differentiating somites of Ndrg2(-/-) embryos, which may partially account for the vertebral defects in Ndrg2 mutants. Nevertheless, Bmp/Smad signaling activity was elevated in the differentiating somites of Ndrg2(-/-) embryos. Collectively, our findings unveiled Ndrg2 as a novel regulator of vertebral specification in differentiating somites.

p38α MAPK Regulates Lineage Commitment and OPG Synthesis of Bone Marrow Stromal Cells to Prevent Bone Loss under Physiological and Pathological Conditions.

Summary Bone marrow-derived mesenchymal stromal cells (BM-MSCs) are capable of differentiating into osteoblasts, chondrocytes, and adipocytes. Skewed differentiation of BM-MSCs contributes to the pathogenesis of osteoporosis. Yet how BM-MSC lineage commitment is regulated remains unclear. We show that ablation of p38α in Prx1+ BM-MSCs produced osteoporotic phenotypes, growth plate defects, and increased bone marrow fat, secondary to biased BM-MSC differentiation from osteoblast/chondrocyte to adipocyte and increased osteoclastogenesis and bone resorption. p38α regulates BM-MSC osteogenic commitment through TAK1-NF-κB signaling and osteoclastogenesis through osteoprotegerin (OPG) production by BM-MSCs. Estrogen activates p38α to maintain OPG expression in BM-MSCs to preserve the bone. Ablation of p38α in BM-MSCs positive for Dermo1, a later BM-MSC marker, only affected osteogenic differentiation. Thus, p38α mitogen-activated protein kinase (MAPK) in Prx1+ BM-MSCs acts to preserve the bone by promoting osteogenic lineage commitment and sustaining OPG production. This study thus unravels previously unidentified roles for p38α MAPK in skeletal development and bone remodeling.

Overexpression of Wnt5a in mouse epidermis causes no psoriasis phenotype but an impairment of hair follicle anagen development.

Increased Wnt5a expression has been observed in psoriatic plaques. However, whether Wnt5a overexpression directly causes psoriasis is unknown. In this study, we generated transgenic (TG) mice with epidermal Wnt5a overexpression under the control of the human K14 promoter. The skin of Wnt5a TG mice was not psoriatic, but characterized with normal proliferation and homeostasis of epidermis. Instead, these TG mice displayed impaired hair follicle transition from telogen to anagen, most likely due to impaired canonical Wnt signalling. These results suggest that increased Wnt5a expression alone is inadequate to induce psoriasis in the skin and possible involvement of Wnt5a in hair follicle cycling.

Misexpression of Pknox2 in Mouse Limb Bud Mesenchyme Perturbs Zeugopod Development and Deltoid Crest Formation

The TALE (Three Amino acid Loop Extension) family consisting of Meis, Pbx and Pknox proteins is a group of transcriptional co-factors with atypical homeodomains that play pivotal roles in limb development. Compared to the in-depth investigations of Meis and Pbx protein functions, the role of Pknox2 in limb development remains unclear. Here, we showed that Pknox2 was mainly expressed in the zeugopod domain of the murine limb at E10.5 and E11.5. Misexpression of Pknox2 in the limb bud mesenchyme of transgenic mice led to deformities in the zeugopod and forelimb stylopod deltoid crest, but left the autopod and other stylopod skeletons largely intact. These malformations in zeugopod skeletons were recapitulated in mice overexpressing Pknox2 in osteochondroprogenitor cells. Molecular and cellular analyses indicated that the misexpression of Pknox2 in limb bud mesenchyme perturbed the Hox10-11 gene expression profiles, decreased Col2 expression and Bmp/Smad signaling activity in the limb. These results indicated that Pknox2 misexpression affected mesenchymal condensation and early chondrogenic differentiation in the zeugopod skeletons of transgenic embryos, suggesting Pknox2 as a potential regulator of zeugopod and deltoid crest formation.

Transcriptional regulator PrqR plays a negative role in glucose metabolism and oxidative stress acclimation in Synechocystis sp. PCC 6803.

Plant and cyanobacteria can perceive signals from soluble sugar and reactive oxygen species (ROS) and then coordinate gene expression under stress acclimation, but the underlying mechanism remains unclear. In this study, we found that the transcriptional factor PrqR (Slr0895) in Synechocystis can perceive signals from ROS generated after shifting from prolonged darkness with glucose into high-light. The deletion mutant (DprqR) showed increased growth rate and decreased ROS content, whereas the complementary strain (CprqR) restored the growth characteristics, phenotypes and ROS status of WT, thereby establishing PrqR as a negative regulator of ROS.LC/GC-MS-based metabolic profiling also showed active ROS mitigation in DprqR mutant. Further study by qRT-PCR, ChIP-PCR and deletion of both prqR and prqA (DprqR-DprqA mutant) revealed that PrqR exerts this negative regulation of ROS removal by controlling the expression of sodB and prqA (slr0896). Furthermore, PrqR also found to control glucose metabolism by regulating a positive regulator of glucose metabolism, sigE, and its regulons. Results suggest that PrqR was involved in perceiving signals from ROS under physiological condition, as well as in regulating stress removal and glucose metabolism.