Abstract
Recent evidence suggests that guanyl nucleotide binding (G) proteins are involved in receptor-mediated bone resorption and in osteoblastic function, but the nature of the G protein coupled to effectors that are involved in these skeletal effects is unknown. The purposes of this study were to determine (1) whether a G protein mediates activation of phosphoinositide-specific phospholipase C in UMR-106 rat osteosarcoma cells, and (2) whether parathyroid hormone (PTH) and a PTH-like protein (PLP) associated with humoral hypercalcemia of malignancy promote GTP-dependent PIP2 hydrolysis. Addition of GTP (10-4 M) or guanosine 5'-0-(3-thiotriphosphate, GTPγS, 10-5 M) to membranes prepared from UMR-106 cells labeled with [3H]myo-inositol increased both [3H]inositol trisphosphate (IP3) and [3H]inositol bisphosphate (IP2) formation. The increases in [3H]IP2 and [3H]IP3 produced by GTP were 8.6- and 4.3-fold, respectively. GTPγS produced a 17.6- and 11.9-fold increase in [3H]IP2 and [3H[IP3, respectively. The stimulatory effects of GTP and GTPγS were dose dependent (GTP ED50 = 3.9 x 10-6 M; GTPγS ED50 = 2.5 x 10-7 M) and progressive over 10 minutes and required the presence of Mg2+. GTP (10-4 M) and GTPγS (10-5 M) decreased membrane [3H]phosphoinositides concomitantly with increased [3H]IP2 and [3H]IP3. The GDP analog guanosine 5'-O-(2-thiodiphosphate, GDPβS) alone did not alter [3H]IP2 or [3H]IP3 production but at 10-4 M blocks the stimulatory effects of GTP and GTPγS. NaF (3 x 10-2 M) produced a 2.8- and 2.0-fold stimulation of [3H]IP2 and [3H]IP3, respectively. In the presence of 10-4 M GTP, bPTH-(1-34) (1 μg/ml) produced an increase in [3H]IP3 and [3H[IP2 of 23.5 ± 3.0% (p < 0.001) and 14.1 ± 2.5% (p < 0.01) within 2 minutes. hPLP-(1-34)amide (1 μg/ml) produced a 19.8 ± 5.3% (p < 0.05) and 13.2 ± 4.8% (p < 0.05) increase in [3H]IP3 and [3H]IP2. We conclude that UMR-106 membranes possess a G protein-sensitive phosphoinositide-specific phospholipase C. Conceivably, this signal transduction pathway contributes to the skeletal actions of PTH and PLP.
| Original language | English (US) |
|---|---|
| Pages (from-to) | 549-556 |
| Number of pages | 8 |
| Journal | Journal of Bone and Mineral Research |
| Volume | 4 |
| Issue number | 4 |
| State | Published - 1989 |
| Externally published | Yes |
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ASJC Scopus subject areas
- Surgery
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G protein-dependent activation of a phosphoinositide-specific phospholipase C in UMR-106 osteosarcoma cell membranes. / Babich, M.; King, K. L.; Nissenson, R. A.
In: Journal of Bone and Mineral Research, Vol. 4, No. 4, 1989, p. 549-556.Research output: Contribution to journal › Article
}
TY - JOUR
T1 - G protein-dependent activation of a phosphoinositide-specific phospholipase C in UMR-106 osteosarcoma cell membranes
AU - Babich, M.
AU - King, K. L.
AU - Nissenson, R. A.
PY - 1989
Y1 - 1989
N2 - Recent evidence suggests that guanyl nucleotide binding (G) proteins are involved in receptor-mediated bone resorption and in osteoblastic function, but the nature of the G protein coupled to effectors that are involved in these skeletal effects is unknown. The purposes of this study were to determine (1) whether a G protein mediates activation of phosphoinositide-specific phospholipase C in UMR-106 rat osteosarcoma cells, and (2) whether parathyroid hormone (PTH) and a PTH-like protein (PLP) associated with humoral hypercalcemia of malignancy promote GTP-dependent PIP2 hydrolysis. Addition of GTP (10-4 M) or guanosine 5'-0-(3-thiotriphosphate, GTPγS, 10-5 M) to membranes prepared from UMR-106 cells labeled with [3H]myo-inositol increased both [3H]inositol trisphosphate (IP3) and [3H]inositol bisphosphate (IP2) formation. The increases in [3H]IP2 and [3H]IP3 produced by GTP were 8.6- and 4.3-fold, respectively. GTPγS produced a 17.6- and 11.9-fold increase in [3H]IP2 and [3H[IP3, respectively. The stimulatory effects of GTP and GTPγS were dose dependent (GTP ED50 = 3.9 x 10-6 M; GTPγS ED50 = 2.5 x 10-7 M) and progressive over 10 minutes and required the presence of Mg2+. GTP (10-4 M) and GTPγS (10-5 M) decreased membrane [3H]phosphoinositides concomitantly with increased [3H]IP2 and [3H]IP3. The GDP analog guanosine 5'-O-(2-thiodiphosphate, GDPβS) alone did not alter [3H]IP2 or [3H]IP3 production but at 10-4 M blocks the stimulatory effects of GTP and GTPγS. NaF (3 x 10-2 M) produced a 2.8- and 2.0-fold stimulation of [3H]IP2 and [3H]IP3, respectively. In the presence of 10-4 M GTP, bPTH-(1-34) (1 μg/ml) produced an increase in [3H]IP3 and [3H[IP2 of 23.5 ± 3.0% (p < 0.001) and 14.1 ± 2.5% (p < 0.01) within 2 minutes. hPLP-(1-34)amide (1 μg/ml) produced a 19.8 ± 5.3% (p < 0.05) and 13.2 ± 4.8% (p < 0.05) increase in [3H]IP3 and [3H]IP2. We conclude that UMR-106 membranes possess a G protein-sensitive phosphoinositide-specific phospholipase C. Conceivably, this signal transduction pathway contributes to the skeletal actions of PTH and PLP.
AB - Recent evidence suggests that guanyl nucleotide binding (G) proteins are involved in receptor-mediated bone resorption and in osteoblastic function, but the nature of the G protein coupled to effectors that are involved in these skeletal effects is unknown. The purposes of this study were to determine (1) whether a G protein mediates activation of phosphoinositide-specific phospholipase C in UMR-106 rat osteosarcoma cells, and (2) whether parathyroid hormone (PTH) and a PTH-like protein (PLP) associated with humoral hypercalcemia of malignancy promote GTP-dependent PIP2 hydrolysis. Addition of GTP (10-4 M) or guanosine 5'-0-(3-thiotriphosphate, GTPγS, 10-5 M) to membranes prepared from UMR-106 cells labeled with [3H]myo-inositol increased both [3H]inositol trisphosphate (IP3) and [3H]inositol bisphosphate (IP2) formation. The increases in [3H]IP2 and [3H]IP3 produced by GTP were 8.6- and 4.3-fold, respectively. GTPγS produced a 17.6- and 11.9-fold increase in [3H]IP2 and [3H[IP3, respectively. The stimulatory effects of GTP and GTPγS were dose dependent (GTP ED50 = 3.9 x 10-6 M; GTPγS ED50 = 2.5 x 10-7 M) and progressive over 10 minutes and required the presence of Mg2+. GTP (10-4 M) and GTPγS (10-5 M) decreased membrane [3H]phosphoinositides concomitantly with increased [3H]IP2 and [3H]IP3. The GDP analog guanosine 5'-O-(2-thiodiphosphate, GDPβS) alone did not alter [3H]IP2 or [3H]IP3 production but at 10-4 M blocks the stimulatory effects of GTP and GTPγS. NaF (3 x 10-2 M) produced a 2.8- and 2.0-fold stimulation of [3H]IP2 and [3H]IP3, respectively. In the presence of 10-4 M GTP, bPTH-(1-34) (1 μg/ml) produced an increase in [3H]IP3 and [3H[IP2 of 23.5 ± 3.0% (p < 0.001) and 14.1 ± 2.5% (p < 0.01) within 2 minutes. hPLP-(1-34)amide (1 μg/ml) produced a 19.8 ± 5.3% (p < 0.05) and 13.2 ± 4.8% (p < 0.05) increase in [3H]IP3 and [3H]IP2. We conclude that UMR-106 membranes possess a G protein-sensitive phosphoinositide-specific phospholipase C. Conceivably, this signal transduction pathway contributes to the skeletal actions of PTH and PLP.
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M3 - Article
C2 - 2554686
AN - SCOPUS:0024317255
VL - 4
SP - 549
EP - 556
JO - Journal of Bone and Mineral Research
JF - Journal of Bone and Mineral Research
SN - 0884-0431
IS - 4
ER -