Objective: To investigate the role of transmembrane protein (TMP) 21 in human thyroid cancer.
Methods: The recombinant expression vector pcDNA3.1 (+)-TMP21 and specific small interfering RNAs (siRNA) against TMP21 were transfected into a papillary thyroid cancer cell line (TPC1). After transfection, the expression of TMP21 was confirmed by quantitative real-time polymerase chain reaction (qRT-PCR) and Western blotting. Moreover, cell viability and apoptosis rate were respectively determined by 3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide (MTT) colorimetric assay and flow cytometry (FCM). Additionally, Western blotting was performed to analyze the adenosine monophosphate (AMP)-activated protein kinase (AMPK)/mammalian target of rapamycin (mTOR) pathways associated protein (P-AMPKα(Thr172), P-mTOR(Ser2448), light chain (LC)-II/LC3-I, and P-S6K(Thr389)) after pre-treatment with AMPK inhibitor, compound C (Com C) and siTMP21.
Results: The TMP21 protein level and cell viability were significantly higher, but apoptotic rate was significantly lower by transfection with pcDNA3.1-TMP21 than those in control group (P < 0.05), and reverse results were obtained by transfection with siTMP21. However, qRT-PCR showed different results due to the feedback inhibition of mRNA. Besides, silencing of TMP21 significantly reduced the levels of P-mTOR(Ser2448) and P-S6K(Thr389) (P < 0.05), but significantly increased the levels of P-AMPKα(Thr172) and LC3-II/LC3-I compared with the control group (P < 0.01). Whereas, the levels of P-AMPKα(Thr172) and LC3-II/LC3-I were significantly decreased by Com C compared with the control group (P < 0.05).
Conclusion: TMP21 modulates cell growth in TPC1 cells by inducing autophagy, which may be associated with activation of AMPK/mTOR pathway.
Keywords: AMPK/mTOR; Thyroid cancer; autophagy; transmembrane protein 21.