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BackgroundTetratricopeptide 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.MethodsBreast 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.ResultsTropomyosin 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.ConclusionSince 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.

Background. The role of disulfidptosis-related lncRNAs remains unclear in lung adenocarcinoma. Methods. Analysis in R software was conducted using different R packages, which are based on the public data from The Cancer Genome Atlas (TCGA) database. The transwell assay was used to evaluate the invasion and migration abilities of lung cancer cells. Results. In our study, we identified 1401 lncRNAs significantly correlated with disulfidptosis-related genes (|Cor| > 0.3 and P < 0.05 ). Then, we constructed a prognosis model consisting of 11 disulfidptosis-related lncRNAs, including AL133445.2, AL442125.1, AC091132.2, AC090948.1, AC020765.2, CASC8, AL606834.1, LINC00707, OGFRP1, U91328.1, and GASAL1. This prognosis model has satisfactory prediction performance. Also, the risk score and clinical information were combined to develop a nomogram. Analyses of biological enrichment and immune-related data were used to identify underlying differences between patients at high-risk and low-risk groups. Moreover, we noticed that the immunotherapy nonresponders have higher risk scores. Meanwhile, patients at a high risk responded more strongly to docetaxel, paclitaxel, and vinblastine. Furthermore, further analysis of the model lncRNA OGFRP1 was conducted, including clinical, immune infiltration, biological enrichment analysis, and a transwell assay. We discovered that by inhibiting OGFRP1, the invasion and migration abilities of lung cancer cells could be remarkably hindered. Conclusion. The results of our study can provide directions for future research in the relevant areas. Moreover, the prognosis signature we identified has the potential for clinical application.

Index 3.4.2 The regulation of TTC9 protein level by other steroid hormones 3.4 The expression of TTC9 in breast cancer cell lines 118 3.5 TTC9 is hormonally regulated in MCF-7 cells 3.6 TTC9 is over-expressed in breast cancer tissues compared with the adjacent normal tissues 122 3.7 Regulation of TTC9 expression in MCF-7 cells 127 3.7.1 TTC9 is serum and growth factors inducible 127 3.7.2 Growth factor-induced TTC9 expression is via the activation of ERK1/2 signaling pathway 3.7.3 The regulations by growth factors and estrogen on TTC9 protein level are mediated through separate pathways 3.8 Subcellular localization of TTC9 3.9 Identification of TTC9 transcript and protein in human tissues 3.10 Detection of TTC9 protein in mice and rats tissues 3.11 TTC9 is also hormonally regulated in mouse uterus 8 3.12 TTC9 interacting proteins 3.13 TTC9 can interact with cellular Tm5NM-1 3.14 TTC9 interacts with Tm5NM-l in mammalian cells 3.15 The linker fragment between the first two TPR domains of TTC9 is important for the interaction with Tm5NM-l ATTENTION: The Singapore Copyright Act applies to the use of this document. Nanyang Technological University Library Index 3.16 TTC9 was expressed at a higher level in adherent cells compared with that in suspension cells 154 DISCUSSION 156 4.1 Sequence characteristics of TTC9 157 4.1.1 mRNA size of TTC9 transcript 157 4.1.2 Characteristics of TTC9 protein 158 4.1.2.1 Protein length of TTC9 158 4.1.2.2 TTC9 is mainly localized in endoplasmic reticulum 159 4.1.2.3 Secondary structure of TTC9 protein 160 4.1.3 TTC9 is located on human chromosome 14 4.2 The involvement of TTC9 in hormone signaling 4.2.1 In ABC28 cells, TTC9 is up-regulated by progesterone through PR 4.2.2 TTC9 is down-regulated by E 2 in MCF-7 cells 4.2.3 TTC9 is involved in MAPK signaling pathways 4.2.3.1 Growth factors increased TTC9 expression 4.2.3.2 TTC9 is differentially regulated by different MAPK signaling pathways 4.2.4 TTC9 is hormonally regulated in vivo 4.2.5 Estrogen and growth factors regulate TTC9 expression through different pathways ATTENTION: The Singapore Copyright Act applies to the use of this document. Nanyang Technological University Library Index 4.3 Identification of TTC9 domains involved in the interaction with Tm5NM-l 4.3.1 The first amino acids of TTC9 is important for the interaction 4.3.2 The second and third TPR domains of TTC9 shows inhibitory effect on the interaction between TTC9 and Tm5NM-1 4.3.3 The first TPR domain and the linker region between the first two TPR domains are important for the interaction 4.4 TTC9 and cytoskeleton organization 4.5 Conclusions 4.