Limits...
Nontranscriptional modulation of intracellular Ca2+ signaling by ligand stimulated thyroid hormone receptor.

Saelim N, John LM, Wu J, Park JS, Bai Y, Camacho P, Lechleiter JD - J. Cell Biol. (2004)

Bottom Line: Coexpression of TRbetaA1 with retinoid X receptor did not enhance regulation.Both xTRbetaA1 and the homologous shortened form of rat TRalpha1 (rTRalphaDeltaF1) localized to the mitochondria and increased O2 consumption, whereas the full-length rat TRalpha1 did neither.We conclude that T3-bound mitochondrial targeted TRs acutely modulate IP3-mediated Ca2+ signaling by increasing mitochondrial metabolism independently of transcriptional activity.

View Article: PubMed Central - PubMed

Affiliation: Department of Cellular and Structural Biology, University of Texas Health Science Center at San Antonio, 78229, USA.

ABSTRACT
Thyroid hormone 3,5,3'-tri-iodothyronine (T3) binds and activates thyroid hormone receptors (TRs). Here, we present evidence for a nontranscriptional regulation of Ca2+ signaling by T3-bound TRs. Treatment of Xenopus thyroid hormone receptor beta subtype A1 (xTRbetaA1) expressing oocytes with T3 for 10 min increased inositol 1,4,5-trisphosphate (IP3)-mediated Ca2+ wave periodicity. Coexpression of TRbetaA1 with retinoid X receptor did not enhance regulation. Deletion of the DNA binding domain and the nuclear localization signal of the TRbetaA1 eliminated transcriptional activity but did not affect the ability to regulate Ca2+ signaling. T3-bound TRbetaA1 regulation of Ca2+ signaling could be inhibited by ruthenium red treatment, suggesting that mitochondrial Ca2+ uptake was required for the mechanism of action. Both xTRbetaA1 and the homologous shortened form of rat TRalpha1 (rTRalphaDeltaF1) localized to the mitochondria and increased O2 consumption, whereas the full-length rat TRalpha1 did neither. Furthermore, only T3-bound xTRbetaA1 and rTRalphaDeltaF1 affected Ca2+ wave activity. We conclude that T3-bound mitochondrial targeted TRs acutely modulate IP3-mediated Ca2+ signaling by increasing mitochondrial metabolism independently of transcriptional activity.

Show MeSH

Related in: MedlinePlus

The pBOX and NLSs of TRβA1 are not required for the acute regulation of Ca2+ signaling. (a) Schematic figure depicting the position of the pBOX deletion in the DBD and the NLS modification within TRβA1. (b) Spatial-temporal stack of IP3-induced Ca2+ wave activity in control oocytes compared with oocytes expressing TR mutants ΔpBox-NLS and ΔNLS. Oocytes expressing the TR mutants were incubated with T3 (100 nM) 10–15 min before IP3 (∼300 nM) injections. (c) Western blot analysis confirming comparable levels of protein expression for both wild-type and mutant TRβA1. (d) Histogram of the average Ca2+ wave periods for each group of oocytes (n values are in parentheses). Statistic significance over control oocytes is indicated by the asterisks (**; ANOVA single factor, P < 0.0001).
© Copyright Policy
Related In: Results  -  Collection


getmorefigures.php?uid=PMC2172460&req=5

fig4: The pBOX and NLSs of TRβA1 are not required for the acute regulation of Ca2+ signaling. (a) Schematic figure depicting the position of the pBOX deletion in the DBD and the NLS modification within TRβA1. (b) Spatial-temporal stack of IP3-induced Ca2+ wave activity in control oocytes compared with oocytes expressing TR mutants ΔpBox-NLS and ΔNLS. Oocytes expressing the TR mutants were incubated with T3 (100 nM) 10–15 min before IP3 (∼300 nM) injections. (c) Western blot analysis confirming comparable levels of protein expression for both wild-type and mutant TRβA1. (d) Histogram of the average Ca2+ wave periods for each group of oocytes (n values are in parentheses). Statistic significance over control oocytes is indicated by the asterisks (**; ANOVA single factor, P < 0.0001).

Mentions: Subsequently, we tested whether the transcriptionally inactive xTRβA1 mutants could still acutely regulate Ca2+ signaling. Oocytes were injected with xTRβA1 mRNA or its mutants and protein expression levels were confirmed using Western analysis 2–3 d after injection (Fig. 4 c). Oocytes expressing xTRβA1 or the mutants were exposed to T3 (100 nM) 10 min before injection with IP3 (∼300 nM). Ca2+ activity was confocally imaged, as described above. The average Ca2+ interwave period for the control group (water-injected oocytes) was 6.6 ± 0.20 s (n = 70), which was significantly shorter than that in the xTRβA1 expressing oocytes (8.40 ± 0.30 s, n = 40; ANOVA single factor, P < 0.0001; Fig. 4 b, d; Fig. 1. More importantly, regulation of the Ca2+ wave period in oocytes expressing either the single mutant xTRβA1-ΔNLS (9.6 ± 0.48 s, n = 24) or the double mutant, xTRβA1ΔpBox-NLS (8.4 ± 0.28 s; n = 24) was indistinguishable from oocytes expressing wild-type xTRβA1 (Fig. 4, b and d). We conclude from these data that neither the pBOX nor the NLS of TRβA1 is required for acute regulation of Ca2+ signaling.


Nontranscriptional modulation of intracellular Ca2+ signaling by ligand stimulated thyroid hormone receptor.

Saelim N, John LM, Wu J, Park JS, Bai Y, Camacho P, Lechleiter JD - J. Cell Biol. (2004)

The pBOX and NLSs of TRβA1 are not required for the acute regulation of Ca2+ signaling. (a) Schematic figure depicting the position of the pBOX deletion in the DBD and the NLS modification within TRβA1. (b) Spatial-temporal stack of IP3-induced Ca2+ wave activity in control oocytes compared with oocytes expressing TR mutants ΔpBox-NLS and ΔNLS. Oocytes expressing the TR mutants were incubated with T3 (100 nM) 10–15 min before IP3 (∼300 nM) injections. (c) Western blot analysis confirming comparable levels of protein expression for both wild-type and mutant TRβA1. (d) Histogram of the average Ca2+ wave periods for each group of oocytes (n values are in parentheses). Statistic significance over control oocytes is indicated by the asterisks (**; ANOVA single factor, P < 0.0001).
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2172460&req=5

fig4: The pBOX and NLSs of TRβA1 are not required for the acute regulation of Ca2+ signaling. (a) Schematic figure depicting the position of the pBOX deletion in the DBD and the NLS modification within TRβA1. (b) Spatial-temporal stack of IP3-induced Ca2+ wave activity in control oocytes compared with oocytes expressing TR mutants ΔpBox-NLS and ΔNLS. Oocytes expressing the TR mutants were incubated with T3 (100 nM) 10–15 min before IP3 (∼300 nM) injections. (c) Western blot analysis confirming comparable levels of protein expression for both wild-type and mutant TRβA1. (d) Histogram of the average Ca2+ wave periods for each group of oocytes (n values are in parentheses). Statistic significance over control oocytes is indicated by the asterisks (**; ANOVA single factor, P < 0.0001).
Mentions: Subsequently, we tested whether the transcriptionally inactive xTRβA1 mutants could still acutely regulate Ca2+ signaling. Oocytes were injected with xTRβA1 mRNA or its mutants and protein expression levels were confirmed using Western analysis 2–3 d after injection (Fig. 4 c). Oocytes expressing xTRβA1 or the mutants were exposed to T3 (100 nM) 10 min before injection with IP3 (∼300 nM). Ca2+ activity was confocally imaged, as described above. The average Ca2+ interwave period for the control group (water-injected oocytes) was 6.6 ± 0.20 s (n = 70), which was significantly shorter than that in the xTRβA1 expressing oocytes (8.40 ± 0.30 s, n = 40; ANOVA single factor, P < 0.0001; Fig. 4 b, d; Fig. 1. More importantly, regulation of the Ca2+ wave period in oocytes expressing either the single mutant xTRβA1-ΔNLS (9.6 ± 0.48 s, n = 24) or the double mutant, xTRβA1ΔpBox-NLS (8.4 ± 0.28 s; n = 24) was indistinguishable from oocytes expressing wild-type xTRβA1 (Fig. 4, b and d). We conclude from these data that neither the pBOX nor the NLS of TRβA1 is required for acute regulation of Ca2+ signaling.

Bottom Line: Coexpression of TRbetaA1 with retinoid X receptor did not enhance regulation.Both xTRbetaA1 and the homologous shortened form of rat TRalpha1 (rTRalphaDeltaF1) localized to the mitochondria and increased O2 consumption, whereas the full-length rat TRalpha1 did neither.We conclude that T3-bound mitochondrial targeted TRs acutely modulate IP3-mediated Ca2+ signaling by increasing mitochondrial metabolism independently of transcriptional activity.

View Article: PubMed Central - PubMed

Affiliation: Department of Cellular and Structural Biology, University of Texas Health Science Center at San Antonio, 78229, USA.

ABSTRACT
Thyroid hormone 3,5,3'-tri-iodothyronine (T3) binds and activates thyroid hormone receptors (TRs). Here, we present evidence for a nontranscriptional regulation of Ca2+ signaling by T3-bound TRs. Treatment of Xenopus thyroid hormone receptor beta subtype A1 (xTRbetaA1) expressing oocytes with T3 for 10 min increased inositol 1,4,5-trisphosphate (IP3)-mediated Ca2+ wave periodicity. Coexpression of TRbetaA1 with retinoid X receptor did not enhance regulation. Deletion of the DNA binding domain and the nuclear localization signal of the TRbetaA1 eliminated transcriptional activity but did not affect the ability to regulate Ca2+ signaling. T3-bound TRbetaA1 regulation of Ca2+ signaling could be inhibited by ruthenium red treatment, suggesting that mitochondrial Ca2+ uptake was required for the mechanism of action. Both xTRbetaA1 and the homologous shortened form of rat TRalpha1 (rTRalphaDeltaF1) localized to the mitochondria and increased O2 consumption, whereas the full-length rat TRalpha1 did neither. Furthermore, only T3-bound xTRbetaA1 and rTRalphaDeltaF1 affected Ca2+ wave activity. We conclude that T3-bound mitochondrial targeted TRs acutely modulate IP3-mediated Ca2+ signaling by increasing mitochondrial metabolism independently of transcriptional activity.

Show MeSH
Related in: MedlinePlus