BMP signaling balances proliferation and differentiation of muscle satellite cell descendants

<p>Abstract</p> <p>Background</p> <p>The capacity of muscle to grow or to regenerate after damage is provided by adult stem cells, so called satellite cells, which are located under the basement lamina of each myofiber. Upon activation satellite cells enter the cell cycle, proliferate and differentiate into myoblasts, which fuse to injured myofibers or form new fibers. These processes are tightly controlled by many growth factors.</p> <p>Results</p> <p>Here we investigate the role of bone morphogenetic proteins (BMPs) during satellite cell differentiation. Unlike the myogenic C2C12 cell line, primary satellite cells do not differentiate into osteoblasts upon BMP signaling. Instead BMP signaling inhibits myogenic differentiation of primary satellite cells <it>ex vivo</it>. In contrast, inhibition of BMP signaling results in cell cycle exit, followed by enhanced myoblast differentiation and myotube formation. Using an <it>in vivo </it>trauma model we demonstrate that satellite cells respond to BMP signals during the regeneration process. Interestingly, we found the BMP inhibitor <it>Chordin </it>upregulated in primary satellite cell cultures and in regenerating muscles. In both systems <it>Chordin </it>expression follows that of Myogenin, a marker for cells committed to differentiation.</p> <p>Conclusion</p> <p>Our data indicate that BMP signaling plays a critical role in balancing proliferation and differentiation of activated satellite cells and their descendants. Initially, BMP signals maintain satellite cells descendants in a proliferating state thereby expanding cell numbers. After cells are committed to differentiate they upregulate the expression of the BMP inhibitor <it>Chordin </it>thereby supporting terminal differentiation and myotube formation in a negative feedback mechanism.</p

Prdm5 Regulates Collagen Gene Transcription by Association with RNA Polymerase II in Developing Bone

PRDM family members are transcriptional regulators involved in tissue specific differentiation. PRDM5 has been reported to predominantly repress transcription, but a characterization of its molecular functions in a relevant biological context is lacking. We demonstrate here that Prdm5 is highly expressed in developing bones; and, by genome-wide mapping of Prdm5 occupancy in pre-osteoblastic cells, we uncover a novel and unique role for Prdm5 in targeting all mouse collagen genes as well as several SLRP proteoglycan genes. In particular, we show that Prdm5 controls both Collagen I transcription and fibrillogenesis by binding inside the Col1a1 gene body and maintaining RNA polymerase II occupancy. In vivo, Prdm5 loss results in delayed ossification involving a pronounced impairment in the assembly of fibrillar collagens. Collectively, our results define a novel role for Prdm5 in sustaining the transcriptional program necessary to the proper assembly of osteoblastic extracellular matrix

Hoxa11 and Hoxd11 Regulate Chondrocyte Differentiation Upstream of Runx2 and Shox2 in Mice

<div><p>During limb development, posterior <em>Hox</em> genes of the <em>Hoxa</em>- and <em>Hoxd</em> cluster provide positional information along the limb axis. Here we report a new function for Hoxa11 and Hoxd11 in regulating the early steps of chondrocyte differentiation. We analyzed forelimbs of <em>Hoxa11^−/−;d11^−/−</em> and <em>Ulnaless</em> mice, which are characterized by specifically shortened zeugopods. By detailed morphological and molecular analyses, we show that loss of Hoxa11 and Hoxd11 in the ulna of both mutants leads to an arrest of chondrocyte differentiation at a step before the separation into round and columnar cells takes place. Furthermore, we demonstrate that Hoxa11 and Hoxd11 act upstream of <em>Runx2</em> and <em>Shox2</em>, two key regulators of chondrocyte differentiation. We hypothesize that Runx2 activates <em>Shox2</em> in early chondrocytes, which at later stages induces <em>Runx2</em> expression to regulate hypertrophic differentiation. These results give insight into mechanisms by which positional information might be translated into a specific bone pattern.</p> </div

Atoh8 acts as a regulator of chondrocyte proliferation and differentiation in endochondral bones.

Atonal homolog 8 (Atoh8) is a transcription factor of the basic helix-loop-helix (bHLH) protein family, which is expressed in the cartilaginous elements of endochondral bones. To analyze its function during chondrogenesis we deleted Atoh8 in mice using a chondrocyte- (Atoh8flox/flox;Col2a1-Cre) and a germline- (Atoh8flox/flox;Prx1-Crefemale) specific Cre allele. In both strains, Atoh8 deletion leads to a reduced skeletal size of the axial and appendicular bones, but the stages of phenotypic manifestations differ. While we observed obviously shortened bones in Atoh8flox/flox;Col2a1-Cre mice only postnatally, the bones of Atoh8flox/flox;Prx1-Crefemale mice are characterized by a reduced bone length already at prenatal stages. Detailed histological and molecular investigations revealed reduced zones of proliferating and hypertrophic chondrocytes. In addition, Atoh8 deletion identified Atoh8 as a positive regulator of chondrocyte proliferation. As increased Atoh8 expression is found in the region of prehypertrophic chondrocytes where the expression of Ihh, a main regulator of chondrocyte proliferation and differentiation, is induced, we investigated a potential interaction of Atoh8 function and Ihh signaling. By activating Ihh signaling with Purmorphamine we demonstrate that Atoh8 regulates chondrocyte proliferation in parallel or downstream of Ihh signaling while it acts on the onset of hypertrophy upstream of Ihh likely by modulating Ihh expression levels

CCN1 (CYR61) and CCN3 (NOV) signaling drives human trophoblast cells into senescence and stimulates migration properties

<p>During placental development, continuous invasion of trophoblasts into the maternal compartment depends on the support of proliferating extravillous trophoblasts (EVTs). Unlike tumor cells, EVTs escape from the cell cycle before invasion into the decidua and spiral arteries. This study focused on the regulation properties of glycosylated and non-glycosylated matricellular CCN1 and CCN3, primarily for proliferation control in the benign SGHPL-5 trophoblast cell line, which originates from the first-trimester placenta. Treating SGHPL-5 trophoblast cells with the glycosylated forms of recombinant CCN1 and CCN3 decreased cell proliferation by bringing about G0/G1 cell cycle arrest, which was accompanied by the upregulation of activated Notch-1 and its target gene p21. Interestingly, both CCN proteins increased senescence-associated β-galactosidase activity and the expression of the senescence marker p16. The migration capability of SGHPL-5 cells was mostly enhanced in response to CCN1 and CCN3, by the activation of FAK and Akt kinase but not by the activation of ERK1/2. In summary, both CCN proteins play a key role in regulating trophoblast cell differentiation by inducing senescence and enhancing migration properties. Reduced levels of CCN1 and CCN3, as found in early-onset preeclampsia, could contribute to a shift from invasive to proliferative EVTs and may explain their shallow invasion properties in this disease.</p

Shox2 acts downstream of both Hoxa11 and Hoxd11 as well as of Runx2.

<p><i>Shox2 in situ</i> hybridization on E14.5 (A–D) and E16.5 (E–H) control (A, E), <i>Hoxa11^−/−;d11^−/−</i> (B, F), <i>Ulnaless</i> (C, G) and <i>Runx2^−/−;Runx3^−/−</i> (D, H) forelimbs revealed the absence of its expression in the ulna of both <i>Hox</i> mutants (B, C, F, G, red arrows) and in ulna and radius of <i>Runx2^−/−;Runx3^−/−</i> mutant mice (D, H, red arrows). However, <i>Shox2</i> expression was detected in other regions of the analyzed forelimbs (B, C, D, G, yellow arrows). 80x magnification (A–C, E–H), 100x magnification (D); R = radius, U = ulna.</p

Chondrocyte differentiation is blocked prior to the separation into columnar and round chondrocytes.

<p>Sections of E14.5 forelimbs of control (A, D, G), <i>Hoxa11^−/−;d11^−/−</i> (B, E, H) and <i>Ulnaless</i> embryos (C, F, I) were hybridized with antisense riboprobes for <i>Col2a1</i> (A–C), <i>Ihh</i> (D–F) and <i>Ptch1</i> (G–I). At E14.5, <i>Col2a1</i> is expressed in all chondrocytes of ulna and radius of <i>Hoxa11^−/−;d11^−/−</i> and <i>Ulnaless</i> mutants similar as in control limbs (A–C). In contrast, <i>Ihh</i>, which is expressed in early hypertrophic chondrocytes in E14.5 control limbs (D, arrow), is not expressed in the ulna of both mutants (E, F). <i>Ptch1</i> is strongly expressed in wild type chondrocytes adjacent to the <i>Ihh</i> expression domain (G, yellow arrow), but only basal expression levels are detected in chondrocytes of the ulna of <i>Hoxa11^−/−;d11^−/−</i> and <i>Ulnaless</i> mice (G–I, red arrows). 80x magnification; R = radius, U = ulna.</p

Hoxa11 and Hoxd11 control early steps of chondrocyte differentiation.

<p>Sections of E14.5 (A–C) and E16.5 (D–F) forelimbs of control (A, D), <i>Hoxa11^−/−;d11^−/−</i> (B, E) and <i>Ulnaless</i> embryos (C, F) were hybridized with antisense riboprobes for <i>Fgfr3</i> (A–C) and <i>Ucma</i> (D–F). High <i>Fgfr3</i> expression (A–C, yellow arrows), which demarcates columnar chondrocytes in control limbs (A), is significantly reduced in the ulna of the <i>Hox</i> mutants (B, C). The observed <i>Fgfr3</i> expression level is comparable with the expression in round chondrocytes of the control mice (A–C, red arrows). <i>Ucma</i>, a marker for round cells (D, arrow), is not expressed in the majority of chondrocytes indicating that the differentiation is blocked before round and columnar chondrocytes are formed (E, F, arrows). 80x magnification; R = radius, U = ulna.</p