Kisspeptin signaling is established as a pathway that can suppress metastasis, but its role in tumors driven by oncogenic HRASG12V had not been fully characterized. This study investigated whether kisspeptin signaling attenuates HRASG12V-induced tumorigenic and metastatic phenotypes in NIH3T3 fibroblast cells and explored the transcriptional regulation of N-cadherin with a focus on SP1-dependent promoter activity.
NIH3T3 cells were engineered to express HRAS G12V, KISS1 (the kisspeptin ligand), and KISS1R (the kisspeptin receptor). The authors evaluated cell proliferation, migration, and invasion in vitro. Reporter assays included an SRF reporter and luciferase constructs driven by the N-cadherin promoter, including deletion constructs to map SP1-responsive regions. Anchorage-independent growth was measured by colony formation assays. N-cadherin expression was assessed by RT-PCR and immunoblotting. Chromatin immunoprecipitation (ChIP) assays were used to measure SP1 binding to the N-cadherin promoter. Finally, in vivo studies tested tumor growth and pulmonary metastasis in mice implanted with the modified NIH3T3 cells. The abstract does not report experimental sample sizes, precise time points, or statistical values.
Kisspeptin signaling reduced NIH3T3 proliferation, migration, and invasion in vitro. Activation of an SRF reporter was observed downstream of kisspeptin signaling. In cells expressing oncogenic HRASG12V, KISS1 expression lowered N-cadherin mRNA and protein levels and suppressed anchorage-independent colony formation.
HRASG12V increased N-cadherin promoter activity. Co-expression of KISS1 reduced both the basal promoter activity and the HRASG12V-stimulated promoter activation. Deletion analysis identified an SP1-responsive region of the N-cadherin promoter required for these regulatory effects. Chromatin immunoprecipitation showed that KISS1 reduced SP1 occupancy at the N-cadherin promoter.
In vivo, expression of KISS1 suppressed HRASG12V-induced tumor growth and pulmonary metastasis. Reintroduction of N-cadherin reversed KISS1’s suppressive influence on tumor growth and metastasis, supporting a causal role for N-cadherin downregulation in the antimetastatic effect of kisspeptin signaling.
Mechanistic experiments indicated that kisspeptin signaling activated an SRF reporter through a signaling cascade involving KISS1R, Gαq/11, p63RhoGEF, and RhoA. Activation of this pathway is consistent with regulation of cytoskeletal and transcriptional programs that can alter cell motility and gene expression. The SRF reporter activation documents downstream functional signaling, while functional assays (migration, invasion, colony formation) showed phenotypic consequences of pathway activation. The abstract does not specify whether pharmacologic inhibitors or genetic knockdowns of individual pathway components were used beyond the constructs explicitly mentioned.
Promoter analysis demonstrated that HRASG12V enhances N-cadherin promoter activity and that KISS1 attenuates both basal and HRASG12V-driven promoter activity. Deletion of the SP1-responsive promoter region abolished regulation by HRASG12V and KISS1, indicating that SP1 sites are required for the observed effects. Chromatin immunoprecipitation confirmed decreased SP1 binding to the N-cadherin promoter in the presence of KISS1, linking kisspeptin signaling to transcriptional repression of N-cadherin via modulation of SP1 occupancy.
In mouse models, KISS1 expression suppressed HRASG12V-driven tumor growth and reduced pulmonary metastasis. The suppressive effect of KISS1 on tumor growth and metastasis was reversed by restoring N-cadherin expression, indicating that downregulation of N-cadherin is necessary for kisspeptin’s antitumor activity in this system. The abstract does not include quantitative tumor burden metrics, numbers of animals, or time-course details; these specifics were not reported in the abstract provided.
The authors conclude that kisspeptin signaling suppresses HRASG12V-induced tumor growth and metastasis in NIH3T3 cells by inhibiting SP1-dependent transcription of N-cadherin. The data support a model in which KISS1–KISS1R activation engages a Gαq/11–p63RhoGEF–RhoA axis and modulates SRF activity while reducing SP1 binding at the N-cadherin promoter, leading to lower N-cadherin expression and diminished tumorigenic and metastatic phenotypes. Details such as experimental sample sizes, statistical tests, and full methodological parameters were not provided in the abstract and would require consultation of the full text for confirmation.