松弛素是松弛素家族RLN2基因编码的多肽激素，调节人体多个系统以适应妊娠过程，是人们研究应用最广泛的多肽激素之一。其家族配体包括RLN1-3和INSL3-6共7个成员，其中Insl6和Insl4的功能尚未清楚，其受体也未知，究其原因可能是现行研究使用的这两者多肽来自化学合成或者原核表达，导致无明显生物活性。本研究拟以演化生物学角度预测Insl6和Insl4的细胞内信号通路和相对应的GPCR受体，另外以可表达内源性Insl6或者Insl4的细胞系来大量表达纯化有生物活性的Insl6和Insl4多肽为基础，利用G蛋白荧光素酶报告系统探测两者在细胞内的G蛋白信号通路，和鉴定出对应的GPCR受体，并进一步对这两个起源于有胎盘类哺乳动物的多肽激素分别对精细胞和胚胎成熟的功能展开深入的揭示，从而解释基因起源对于物种分化形成的重要作用，同时也为开发Insl6和Insl4药物提供生物学机理和应用基础。

Relaxin, the peptide product of RLN2 gene in relaxin family, has been shown to play a key role in the maternal hemodynamic and renal adjustments that accommodate pregnancy. The relaxin family members include seven peptide hormones-RLN1-3, INSL3-6. However, the function and receptor of INSL6 and INSL4 remain uncharacterized. The marketed Insl6 and Insl4 are probably non-active peptides due to chemical synthesis or expression from the bacterial. Here our research bases on evolution analysis of relaxin ligand-receptor relationship and focus on the expression and purification of Insl6 or Insl4 peptide from the special mammalian cell lines which can express the endogens Insl6 or Insl4 very well. Based on the active Insl6 and Insl4 peptide we purified, we will further indentify their intracellular G protein signaling and their cognate GPCR receptor with G protein luciferase reporter assay. Finally we will figure out the cellular mechanism of Insl6 and Insl4 on spermatogenesis and embryogenesis, respectively. Our result will decipher the effect of new gene origin on the species differentiation and also privide the biological basis for pharmacological study of Insl6 and Insl4.