7170

TPM3

tropomyosin 3;TPM3;tropomyosin 3

alpha-tropomyosin, slow skeletal|cytoskeletal tropomyosin TM30|epididymis luminal protein 189|epididymis secretory sperm binding protein Li 82p|heat-stable cytoskeletal protein 30 kDa|tropomyosin alpha-3 chain|tropomyosin gamma|tropomyosin-5

1q21.2

protein-coding

BACKGROUND: Tetratricopeptide repeat domain 9A (TTC9A) protein is a recently identified protein which contains three tetratricopeptide repeats (TPRs) on its C-terminus. In our previous studies, we have shown that TTC9A was a hormonally-regulated gene in breast cancer cells. In this study, we found that TTC9A was over-expressed in breast cancer tissues compared with the adjacent controls (P < 0.00001), suggesting it might be involved in the breast cancer development process. The aim of the current study was to further elucidate the function of TTC9A. METHODS: Breast samples from 25 patients including the malignant breast tissues and the adjacent normal tissues were processed for Southern blot analysis. Yeast-two-hybrid assay, GST pull-down assay and co-immunoprecipitation were used to identify and verify the interaction between TTC9A and other proteins. RESULTS: Tropomyosin Tm5NM-1 was identified as one of the TTC9A partner proteins. The interaction between TTC9A and Tm5NM-1 was further confirmed by GST pull-down assay and co-immunoprecipitation in mammalian cells. TTC9A domains required for the interaction were also characterized in this study. The results suggested that the first TPR domain and the linker fragment between the first two TPR domains of TTC9A were important for the interaction with Tm5NM-1 and the second and the third TPR might play an inhibitory role. CONCLUSION: Since the primary function of tropomyosin is to stabilize actin filament, its interaction with TTC9A may play a role in cell shape and motility. In our previous results, we have found that progesterone-induced TTC9A expression was associated with increased cell motility and cell spreading. We speculate that TTC9A acts as a chaperone protein to facilitate the function of tropomyosins in stabilizing microfilament and it may play a role in cancer cell invasion and metastasis.

High and low molecular weight (LMW) tropomyosin isoforms, by regulation of actin filaments, have a major role in the regulation of cell behaviour. They affect malignant transformation, motility, differentiation, metastasis and cell membrane protein presentation. expression of LMW isoforms from the TPM1 and TPM3 genes have an important role in these effects but the regulation of their expression is unknown. Luciferase assays on a progressively truncated 1.7 kb fragment upstream of the exon 1b translation start site in the TPM1 and TPM3 genes in HEK-293 cells showed upstream activation sequences in TPM1 between -152 and -139 bp and in TPM3 between -154 and -102 bp. The effect of mutating candidate transcription factor binding sites identified an AML1-like transcription factor binding site in TPM1 and a cAMP response element in TPM3. Downstream from the primary activation sequence in TPM1 was a repressor region corresponding to two Sp/KLF family binding GC boxes. Band shift assays confirmed that deletion of these sites altered transcription factor binding and ChIP assays confirmed the presence of AML1 and CREB at the TPM1 and TPM3 activation sequences in the respective promoters. expression of LMW isoforms from TPM1 and TPM3 genes is regulated very differently. This facilitates regulation of the many cell processes involving these proteins. In situations where these proteins have a critical role, such as cancer metastasis, it also facilitates specific intervention.