Evidence for the involvement of adenomatous polyposis coli (APC) protein in maintaining cellular distributions of α3β4 nicotinic receptors

• Published in: Neuroscience letters 2011-02, Vol.489 (2), p.105-109
• Main Authors: Potaros, Tulaya, Phornchirasilp, Srichan, McKay, Susan B., González-Cestari, Tatiana F., Boyd, R. Thomas, McKay, Dennis B.
• Format: Article
• Language: English
• Subjects: Adenomatous Polyposis Coli Protein - genetics; Adenomatous Polyposis Coli Protein - physiology; Adrenal Glands - metabolism; Adrenal medulla; Alpha 3; Animals; APC; Biological and medical sciences; Cattle; Cell Membrane - metabolism; Chromaffin Cells - metabolism; Fundamental and applied biological sciences. Psychology; HEK293 Cells; Humans; Intracellular receptors; Nicotinic receptor; Receptors, Nicotinic - metabolism; Recombinant Proteins - metabolism; RNA, Small Interfering - genetics; siRNA; Vertebrates: nervous system and sense organs
• ISSN: 0304-3940; 1872-7972
• DOI: 10.1016/j.neulet.2010.11.075

▶ BAC and BMα3β4 cells express APC protein. ▶ APC siRNA reduces APC protein levels. ▶ APC siRNA reduces intracellular nAChRs without affecting total number of nAChRs. ▶De novo APC protein synthesis is important for maintenance of nAChRs distribution. ▶ APC protein is involved in the maintenance of cellular distribution of α3β4 nAChRs. Evidence exists supporting the involvement of adenomatous polyposis coli (APC) protein in the assembly of neuronal nicotinic acetylcholine receptors (nAChRs) in the postsynaptic complex. In the following studies, the effects of APC protein on cellular distribution of recombinant α3β4 nAChRs was investigated. RT-PCR and Western blotting techniques established the expression of APC protein both in bovine adrenal chromaffin cells, which express native α3β4* nAChRs, and in a HEK293 cell line expressing recombinant bovine adrenal α3β4 nAChRs (BMα3β4 cells). Transfection of BMα3β4 cells with siRNA to APC, reduced APC protein levels to 52.4% and 61.9% of control values at 24 and 48h after transfection. To investigate the effects of APC on the cellular distribution of α3β4 nAChRs, [3H]epibatidine binding approaches, coupled with APC siRNA treatment, were used. Twenty-four and 48h after APC siRNA transfection, intracellular nAChRs were significantly reduced to 71% and 68% of control, respectively, while the total population of nAChRs were not significantly changed. Given that total cellular nAChRs represent the sum of surface and intracellular nAChRs, these studies support a re-distribution of nAChRs to the plasma membrane with APC siRNA treatment. Treatment of the cells with the protein synthesis inhibitor, puromycin, also caused a significant reduction (55%) in APC protein levels, and produced a similar re-distribution of cellular nAChRs. These studies support the involvement of APC protein in the maintenance of normal cellular distribution of α3β4 nAChRs.