1,25-DIHYDROXYVITAMIN D-3 ATTENUATES ADENYLYL-CYCLASE ACTIVITY IN RAT-THYROID CELLS - REDUCTION OF THYROTROPIN RECEPTOR NUMBER AND INCREASEIN GUANINE-NUCLEOTIDE-BINDING PROTEIN G(I-2)ALPHA
Jp. Berg et al., 1,25-DIHYDROXYVITAMIN D-3 ATTENUATES ADENYLYL-CYCLASE ACTIVITY IN RAT-THYROID CELLS - REDUCTION OF THYROTROPIN RECEPTOR NUMBER AND INCREASEIN GUANINE-NUCLEOTIDE-BINDING PROTEIN G(I-2)ALPHA, Endocrinology, 135(2), 1994, pp. 595-602
1,25-Dihydroxyvitamin D-3 [1,25-(OH)(2)D-3] is the most potent of the
naturally occurring vitamin D metabolites. In rat thyroid FRTL-5 cells
, 1,25-(OH)(2)D-3 attenuated the increase in TSH-stimulated adenylyl c
yclase activity obtained by removing TSH from the culture medium. When
cells were incubated with 1,25-(OH)(2)D-3 (10 nmol/liter; 4 days), th
e binding capacity for specific [I-125]TSH binding decreased from 20.1
+/- 1.8 to 8.8 +/- 1.6 fmol/10(6) cells (mean +/- SEM; n = 4; P < 0.0
1) compared to that in control cells. The K-d did not change (mean +/-
SEM, 0.46 +/- 0.09 vs. 0.25 +/- 0.07 nmol/liter; n = 4; P = NS). West
ern blotting revealed no change in the membrane content of the adenyly
l cyclase (AC) stimulatory guanine nucleotide-binding protein (G-prote
in) alpha-subunit (G(s) alpha) during 1,25-(OH)(2)D-3 treatment. Simil
arly, levels of the AC inhibitory G-protein G(i.3)alpha- and G-protein
beta-subunits were not altered by 1,25-(OH)(2)D-3. However, Western b
lotting with antibodies recognizing both G(i-1)alpha and G(i-2)alpha w
as augmented 4-fold, presumably representing an increase in G(i.2)alph
a only, as G(i-1)alpha messenger RNA (mRNA) was not detected in FRTL-5
cells. 1,25-(OH)(2)D-3 (10 nmol/liter; 4 days) reduced cholera toxin
(10 nmol/liter)-stimulated AC activity to 85% of the control value (P
< 0.05), whereas forskolin (100 mu mol/liter)-stimulated direct activa
tion of AC was inhibited by 39%. The TSH receptor mRNA level correlate
d to the beta-actin mRNA was 2-fold higher in control cells compared t
o that in 1,25-(OH)(2)D-3-treated cells 12 h after TSH removal. Only m
inor alterations in the G(i-3)alpha mRNA/beta-actin mRNA and G(i-3)alp
ha mRNA/beta-actin mRNA ratios were observed during 1,25-(OH)(2)D-3 tr
eatment, whereas G(i-2)alpha mRNA increased 9-fold compared to that in
control cells. No change in the resting intracellular Ca2+ concentrat
ion could be detected after 4 days of 1,25-(OH)(2)D-3 treatment. Our s
tudies show that 1,25-(OH)(2)D-3 attenuates AC activity by reducing th
e TSH receptor number and increasing the level of the AC inhibitory G-
protein G(i-2)alpha in FRTL-5 cells.