Mouse thromboxane A2 receptor: cDNA cloning, expression and northern blot analysis

Area of Science:

• Molecular Biology
• Pharmacology
• Immunology

Background:

• The thromboxane A2 receptor (TP) plays a crucial role in various physiological processes, including platelet aggregation and inflammation.
• Understanding the molecular characteristics and distribution of the TP is essential for developing targeted therapies.

Purpose of the Study:

• To isolate and characterize the cDNA clone for the mouse thromboxane A2 receptor.
• To confirm the functional activity and ligand-binding properties of the cloned receptor.
• To investigate the tissue-specific expression of the thromboxane A2 receptor mRNA in mice.

Main Methods:

• Isolation of a mouse thromboxane A2 receptor cDNA clone from a mouse lung cDNA library.
• Expression of the receptor transcript in Xenopus laevis oocytes to assess functional activity.
• Radioligand binding assays using [3H]S-145 in COS-1 cells transfected with the cDNA.
• Northern blot analysis to determine the distribution of thromboxane A2 receptor mRNA.

Main Results:

• A 1,023 bp open reading frame encoding a 341 amino acid protein was identified.
• Functional assays demonstrated that STA2 and U-46619 induced inward currents in oocytes expressing the receptor.
• Specific binding of [3H]S-145 was observed with a Kd of 3.3 nM, and binding was competitively inhibited by various prostaglandins and thromboxane analogues.
• Northern blot analysis revealed abundant expression of thromboxane A2 receptor mRNA in the thymus, spleen, and lung.

Conclusions:

• The isolated cDNA clone represents the functional mouse thromboxane A2 receptor.
• The receptor exhibits specific binding characteristics for thromboxane analogues.
• Significant expression in immune organs suggests a role for the thromboxane A2 receptor in immune responses.