Recombinant Lactococcus lactis NZ9000 secretes a bioactive kisspeptin that inhibits proliferation and migration of human colon carcinoma HT-29 cells.
BACKGROUND
Proteinaceous bioactive substances and pharmaceuticals are most conveniently administered orally. However, the facing problems are the side effects of proteolytic degradation and denaturation in the gastrointestinal tract. In recent years, lactic acid bacteria (LAB) have been verified to be a promising delivery vector for susceptible functional proteins and drugs. KiSS-1 peptide, a cancer suppressor, plays a critical role in inhibiting cancer metastasis and its activity has been confirmed by direct administration. However, whether this peptide can be functionally expressed in LAB and exert activity on cancer cells, thus providing a potential alternative administration manner in the future, has not been demonstrated.


RESULTS
A recombinant Lactococcus lactis strain NZ9000-401-kiss1 harboring a plasmid containing the gene of the tumor metastasis-inhibiting peptide KiSS1 was constructed. After optimization of the nisin induction conditions, the recombinant strain efficiently secreted KiSS1 with a maximum detectable amount of 27.9 μg/ml in Dulbecco's Modified Eagle medium. The 3-[4,5-dimethylthiazol-2-yl]-2,5 diphenyl tetrazolium bromide and would healing assays, respectively, indicated that the secreted KiSS1 peptide remarkably inhibited HT-29 cell proliferation and migration. Furthermore, the expressed KiSS1 was shown to induce HT-29 cell morphological changes, apoptosis and reduce the expression of matrix metalloproteinase 9 (MMP-9) at both mRNA and protein levels.


CONCLUSIONS
A recombinant L. lactis NZ9000-401-kiss1 successfully expressing the human kiss1 was constructed. The secreted KiSS1 peptide inhibited human HT-29 cells' proliferation and migration probably by invoking, or mediating the cell-apoptosis pathway and by down regulating MMP-9 expression, respectively. Our results suggest that L. lactis is an ideal cell factory for secretory expression of tumor metastasis-inhibiting peptide KiSS1, and the KiSS1-producing L. lactis strain may serve as a new tool for cancer therapy in the future.
Beijing Advanced Innovation Center for Food Nutrition and Human Health, College of Food Science and Nutritional Engineering, China Agricultural University, No. 17 Qinghua East Road, Haidian District, Beijing, 100083, People's Republic of China.|Key Laboratory of Functional Dairy, Department of Food Science and Engineering, College of Food Science and Nutritional Engineering, China Agricultural University, Beijing, 100083, People's Republic of China.
Bo
B
Zhang
Key Laboratory of Functional Dairy, Department of Food Science and Engineering, College of Food Science and Nutritional Engineering, China Agricultural University, Beijing, 100083, People's Republic of China.
Angdi
A
Li
Key Laboratory of Functional Dairy, Department of Food Science and Engineering, College of Food Science and Nutritional Engineering, China Agricultural University, Beijing, 100083, People's Republic of China.
Fanglei
F
Zuo
Beijing Advanced Innovation Center for Food Nutrition and Human Health, College of Food Science and Nutritional Engineering, China Agricultural University, No. 17 Qinghua East Road, Haidian District, Beijing, 100083, People's Republic of China.|Key Laboratory of Functional Dairy, Department of Food Science and Engineering, College of Food Science and Nutritional Engineering, China Agricultural University, Beijing, 100083, People's Republic of China.
Rui
R
Yu
Beijing Advanced Innovation Center for Food Nutrition and Human Health, College of Food Science and Nutritional Engineering, China Agricultural University, No. 17 Qinghua East Road, Haidian District, Beijing, 100083, People's Republic of China.|Key Laboratory of Functional Dairy, Department of Food Science and Engineering, College of Food Science and Nutritional Engineering, China Agricultural University, Beijing, 100083, People's Republic of China.
Zhu
Z
Zeng
College of Horticulture, China Agricultural University, Beijing, 100193, People's Republic of China.
Huiqin
H
Ma
Beijing Advanced Innovation Center for Food Nutrition and Human Health, College of Food Science and Nutritional Engineering, China Agricultural University, No. 17 Qinghua East Road, Haidian District, Beijing, 100083, People's Republic of China. swchen@cau.edu.cn.|Key Laboratory of Functional Dairy, Department of Food Science and Engineering, College of Food Science and Nutritional Engineering, China Agricultural University, Beijing, 100083, People's Republic of China. swchen@cau.edu.cn.
Shangwu
S
Chen
D017209:Apoptosis / Q000187:drug effects; D002465:Cell Movement / Q000187:drug effects; D049109:Cell Proliferation / Q000187:drug effects; D015536:Down-Regulation / Q000187:drug effects; D019073:HT29 Cells; D006801:Humans; D059648:Kisspeptins / Q000235:genetics / Q000378:metabolism / Q000494:pharmacology*; D013294:Lactococcus lactis / Q000235:genetics / Q000254:growth & development / Q000378:metabolism*; D020780:Matrix Metalloproteinase 9 / Q000235:genetics / Q000378:metabolism; D010957:Plasmids / Q000235:genetics / Q000378:metabolism; D043562:Receptors, G-Protein-Coupled / Q000378:metabolism; D000074502:Receptors, Kisspeptin-1; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000302:isolation & purification / Q000494:pharmacology
C118688:KISS1R protein, human; D059648:Kisspeptins; D043562:Receptors, G-Protein-Coupled; D000074502:Receptors, Kisspeptin-1; D011994:Recombinant Proteins; D020780:Matrix Metalloproteinase 9
Delivery vector; Functional protein; KiSS1; Lactococcus lactis