Thyrotropin-releasing Hormone (TRH) Receptor Number Determines the Size of the TRH-responsive Phosphoinositide Pool

Demonstration using controlled expression of TRH receptors by adenovirus mediated gene transfer

Marvin C. Gershengorn, Marcos Heinflink, Daniel R. Nussenzveig, Patricia M. Hinkle, Erik Falck-Pedersen

Research output: Contribution to journalArticle

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Abstract

We use an adenovirus vector, AdCMVmTRHR, to express thyrotropin-releasing hormone (TRH) receptors (TRH-Rs) to determine whether the size of the hormone- responsive phosphoinositide pool in mammalian cells is directly related to receptor number. Infection of HeLa cells with increasing numbers of AdCMVmTRHR caused time-dependent graded expression of TRH-Rs. Measurement of cytoplasmic free Ca2+ in individual cells permitted quantitation of the fraction of cells responsive to TRH. Infection with 100 AdCMVmTRHR particles/cell or more led to TRH responsiveness in ≥90% of HeLa cells. Measurement of prelabeled phosphoinositides hydrolyzed during prolonged TRH stimulation assesses the size of the TRH-responsive pool. In cells infected with AdCMVmTRHR for 24 h, the size of the TRH-responsive phosphoinositide pool increased with increasing TRH-R expression. The TRH-responsive pool also increased with time after infection as the number of TRH-Rs increased. Similar observations were made in GHY and KB cells. These data confirm our previous suggestion (Cubitt, A. B., Geras-Raaka, E., and Gershengorn, M. C. (1990) Biochem. J. 271, 331-336) that there are hormone- responsive and -unresponsive pools of cellular phosphoinositides and that the maximal size of the TRH- responsive pool is directly related to the number of TRH-Rs.

Original languageEnglish (US)
Pages (from-to)6779-6783
Number of pages5
JournalJournal of Biological Chemistry
Volume269
Issue number9
StatePublished - Mar 4 1994

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Thyrotropin Releasing Hormone Receptors
Gene transfer
Thyrotropin-Releasing Hormone
Phosphatidylinositols
Adenoviridae
Demonstrations
Genes
HeLa Cells
Infection
Hormones
KB Cells

ASJC Scopus subject areas

  • Biochemistry

Cite this

Thyrotropin-releasing Hormone (TRH) Receptor Number Determines the Size of the TRH-responsive Phosphoinositide Pool : Demonstration using controlled expression of TRH receptors by adenovirus mediated gene transfer. / Gershengorn, Marvin C.; Heinflink, Marcos; Nussenzveig, Daniel R.; Hinkle, Patricia M.; Falck-Pedersen, Erik.

In: Journal of Biological Chemistry, Vol. 269, No. 9, 04.03.1994, p. 6779-6783.

Research output: Contribution to journalArticle

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abstract = "We use an adenovirus vector, AdCMVmTRHR, to express thyrotropin-releasing hormone (TRH) receptors (TRH-Rs) to determine whether the size of the hormone- responsive phosphoinositide pool in mammalian cells is directly related to receptor number. Infection of HeLa cells with increasing numbers of AdCMVmTRHR caused time-dependent graded expression of TRH-Rs. Measurement of cytoplasmic free Ca2+ in individual cells permitted quantitation of the fraction of cells responsive to TRH. Infection with 100 AdCMVmTRHR particles/cell or more led to TRH responsiveness in ≥90{\%} of HeLa cells. Measurement of prelabeled phosphoinositides hydrolyzed during prolonged TRH stimulation assesses the size of the TRH-responsive pool. In cells infected with AdCMVmTRHR for 24 h, the size of the TRH-responsive phosphoinositide pool increased with increasing TRH-R expression. The TRH-responsive pool also increased with time after infection as the number of TRH-Rs increased. Similar observations were made in GHY and KB cells. These data confirm our previous suggestion (Cubitt, A. B., Geras-Raaka, E., and Gershengorn, M. C. (1990) Biochem. J. 271, 331-336) that there are hormone- responsive and -unresponsive pools of cellular phosphoinositides and that the maximal size of the TRH- responsive pool is directly related to the number of TRH-Rs.",
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