Stimulatory G protein directly regulates hypertrophic differentiation of growth plate cartilage
            <i>in vivo</i>

Baştepe, Murat; Weinstein, Lee S.; Ogata, Naoshi; Kawaguchi, Hiroshi; Jüppner, Harald; Kronenberg, Henry M.; Chung, Ung‐il

doi:10.1073/pnas.0405091101

Cited by 141 publications

(107 citation statements)

References 33 publications

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“…This is also explains why BGsKO mice show no significant changes within the proliferative and hypertrophic zones of the epiphyseal growth plate at later stages. Chondrocyte-specific G s ␣ knock-out and other knock-out models suggest that regulation of the endochondral growth plates is mediated through PTHrP-PPR-G s ␣ pathways (3,4,12,26).…”

Section: Discussionmentioning

confidence: 99%

Deficiency of the G-protein α-Subunit Gsα in Osteoblasts Leads to Differential Effects on Trabecular and Cortical Bone

Sakamoto¹,

Chen²,

Nakamura

et al. 2005

Journal of Biological Chemistry

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The G-protein ␣-subunit G s ␣ is required for the intracellular cAMP responses to hormones and other agonists. G s ␣ is known to mediate the cAMP response to parathyroid hormone and other hormones and cytokines in bone and cartilage. To analyze the in vivo role of G s ␣ signaling in osteoblasts, we developed mice with osteoblast/osteocyte-specific G s ␣ deficiency (BGsKO) by mating G s ␣-floxed mice with collagen I␣1 promoter-Cre recombinase transgenic mice. Early skeletal development was normal in BGsKO mice, because formation of the initial cartilage template and bone collar was unaffected. The chondrocytic zones of the growth plates also appeared normal in BGsKO mice. BGsKO mice had a defect in the formation of the primary spongiosa with reduced immature osteoid (new bone formation) and overall length, which led to reduced trabecular bone volume. In contrast, cortical bone was thickened with narrowing of the bone marrow cavity. This was probably due to decreased cortical bone resorption, because osteoclasts were markedly reduced on the endosteal surface of cortical bone. In addition, the expression of alkaline phosphatase, an early osteoblastic differentiation marker, was normal, whereas the expression of the late osteoblast differentiation markers osteopontin and osteocalcin was reduced, suggesting that the number of mature osteoblasts in bone is reduced. Expression of the osteoclast-stimulating factor receptor activator of NF-B ligand was also reduced. Overall, our findings have similarities to parathyroid hormone null mice and confirm that the differential effects of parathyroid hormone on trabecular and cortical bone are primarily mediated via G s ␣ in osteoblasts. Osteoblast-specific G s ␣ deficiency leads to reduced bone turnover.1 is a ubiquitously expressed G-protein ␣-subunit that couples receptors to adenylyl cyclase and is required for receptor-stimulated cAMP generation (1). cAMP is an important second messenger in osteoblasts for several hormones and other extracellular factors, such as parathyroid hormone (PTH) and prostaglandin E 2 . Both PTH and PTH-related peptide (PTHrP), a paracrine regulator of chondrocyte differentiation in growth plates, activate a common receptor (PTH/PTHrP receptor, PPR), which activates G s ␣ as well as other G-proteins (2-4). PTH stimulates bone formation and bone resorption, resulting in high bone remodeling (5).Different bones are formed by one of two different mechanisms, either intramembranous or endochondral ossification. Intramembranous ossification, which leads to formation of many of the flat bones, occurs when pluripotent mesenchymal cells directly enter an osteoblast lineage and differentiate into osteoblastic cells. Long bones develop by endochondral ossification in which a cartilage template is formed, and chondrocytes undergo hypertrophic differentiation and are then replaced by osteoblasts that form the primary spongiosa, which eventually forms the trabecular bone. Simultaneously, perichondrial cells surrounding the hypertrophic chondrocytes differentia...

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Section: Discussionmentioning

confidence: 99%

Deficiency of the G-protein α-Subunit Gsα in Osteoblasts Leads to Differential Effects on Trabecular and Cortical Bone

Sakamoto¹,

Chen²,

Nakamura

et al. 2005

Journal of Biological Chemistry

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“…15,38 In chondrocytes, PTHrP is regulated by cAMP and transduces signals through the PTH/PTHrP receptor, activating adenylate cyclase for cAMP production. 39,40 We suggest that increased cAMP hydrolysis could cause PTHrP downregulation through a regulatory feedback loop (Fig. 5d).…”

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confidence: 91%

“…(PTH/PTHrP) receptor in chondrocytes and activates Gs-α, thereby stimulating the adenylate 27 cyclase (AC) to produce cAMP. 39 Since PDE3A hydrolyses cAMP, PKA is cAMPdependently regulated and activates CREB, which in turn regulates the PTHLH expression. 40 …”

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confidence: 99%

PDE3A mutations cause autosomal dominant hypertension with brachydactyly

et al. 2015

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All Mendelian hypertension syndromes described to date involve increased sodium reabsorption in the distal nephron. 5 The sole exception is autosomal-dominant hypertension with BDE (HTNB, OMIM #112410), first reported in a Turkish kindred. 2,6 HTNB was linked to chromosome 12p in six unrelated families. 2,7,8 The locus accounts for a ~50 mm Hg mean blood pressure difference at age 50 years. 2 The penetrance is 100% (Fig. 1a). Previously, we reported a rearrangement on chromosome 12p common to all families. 8,9 A linkage study in Chinese hypertensive families without BDE coincided with the HTNB locus, supporting relevance to essential hypertension. 10 Whole-genome sequencing of Turkish family members revealed a heterozygous missense mutation in PDE3A (Gene ID: 5139), a gene encoding a cGMP/cAMP phosphodiesterase with a prominent role in the heart, VSMC, oocytes and platelets. 11 Resequencing of all 48 affected persons in six unrelated families identified six independently clustered heterozygous missense mutations in exon 4 (Fig. 1a, b Supplementary Fig. 1).We detected none of the previously described chromosomal breakpoints on chromosome 12p12.2-12.1, perhaps due to high repetitive content in the breakpoint regions Fig. 2a-c). 4 A haplotype analysis identified a novel recombination that reduced the linkage interval and eliminated an inversion common to all affected individuals in the six families (Fig. 2c). 9 In contrast, the affected mother's haplotype showed co-segregation with the more severe brachydactyly phenotype.PDEs are involved during early stages of osteogenesis. 12 PDE4D mutations have been associated with severe brachydactyly in acrodysostosis. 13,14 In mice, Pde3a was expressed in the developing limbs, consistent with a role during chondrogenesis (Fig. 2d, Supplementary Fig. 3a, b). Chondrogenic downregulation of PTHLH encoding PTHrP was associated with BDE. 15 We also observed PTHLH downregulation in chondrogenically induced fibroblasts from affected persons (Fig. 2e, Supplementary Fig. 3c).We addressed the functional consequences of the identified PDE3A mutations in HeLa cells expressing the six mutations. Forskolin or L-arginine stimulated the adenylate or guanylate cyclases to enhance cellular cAMP or cGMP levels, respectively. 16,17 We detected significantly reduced cAMP levels, consistent with gain-of-function mutations with no change in cGMP levels for the PDE3A mutations ( Supplementary Fig. 4a, b). Three PDE3A isoforms, PDE3A1 (microsomal), PDE3A2 and PDE3A3 (microsomal and cytosolic), have been identified in human myocardium. 18,19 PDE3A3 does not contain the sequence harboring the detected mutations. The predominant isoform in VSMC is PDE3A2. 18,20 To directly elucidate the mutations' effects, we compared the Michaelis-Menten kinetics of cAMPhydrolytic activity for recombinant T445N FLAG-tagged PDE3A1 and PDE3A1-WT and the tagged A2 isoforms purified from transfected cells (Fig. 3a, b, Supplementary Fig. 4d-k). The T445N mutation increased the affinity of both enzyme's isoforms for cAM...

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“…Quantitative real time PCR was performed using primers for Gnas (43), Runx2 (44), Osx (44), collagen I␣1 (ColI␣1) (40), osteopontin (Opn) (44), osteocalcin (Bglap) (45), and Sost (46) according to previously published protocols with mRNA levels normalized relative to ␤-actin expression. Total RNA samples subjected to cDNA synthesis reactions in the absence of reverse transcriptase were included as negative controls.…”

Section: Methodsmentioning

confidence: 99%

Loss of Gsα in the Postnatal Skeleton Leads to Low Bone Mass and a Blunted Response to Anabolic Parathyroid Hormone Therapy

Sinha

Aarnisalo

Chubb

et al. 2016

Journal of Biological Chemistry

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. Surprisingly, in both male and female mice, PTH administration significantly increased osteoblast numbers and bone formation rate in both control and P-G s ␣ OsxKO mice. In mice that express a mutated PTH1R that activates adenylyl cyclase and protein kinase A (PKA) via G s ␣ but not phospholipase C via G q/11 (D/D mice), PTH significantly enhanced bone formation, indicating that phospholipase C activation is not required for increased bone turnover in response to PTH. Therefore, although the anabolic effect of intermittent PTH treatment on trabecular bone volume is blunted by deletion of G s ␣ in osteoblasts, PTH can stimulate osteoblast differentiation and bone formation. Together these findings suggest that alternative signaling pathways beyond G s ␣ and G q/11 act downstream of PTH on osteoblast differentiation.

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Stimulatory G protein directly regulates hypertrophic differentiation of growth plate cartilage in vivo

Cited by 141 publications

References 33 publications

Deficiency of the G-protein α-Subunit Gsα in Osteoblasts Leads to Differential Effects on Trabecular and Cortical Bone

Deficiency of the G-protein α-Subunit Gsα in Osteoblasts Leads to Differential Effects on Trabecular and Cortical Bone

PDE3A mutations cause autosomal dominant hypertension with brachydactyly

Loss of Gsα in the Postnatal Skeleton Leads to Low Bone Mass and a Blunted Response to Anabolic Parathyroid Hormone Therapy

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