Molecular structure responsible for nuclear translocation of phospholipase C-zeta

Abstract

Phospholipase C-zeta (PLCζ) is a strong candidate of the mammalian sperm factor that induces IP₃-mediated Ca²⁺ oscillations and subsequent embryonic development. PLCζ consists of 4 EF-hand domains (EF1-4) in the N terminus, X and Y catalytic domains, and C2 domain in the C terminus. PLCζ expressed by injection of cRNA into mouse eggs causes fertilization-like Ca²⁺ oscillations, and then it is accumulated into the formed pronucleus as the sperm factor is. The nuclear translocation ability (NTA) was investigated by expressing PLCζ mutants tagged with a fluorescent protein by RNA injection into eggs or 1-cell embryos. Point mutation analysis revealed a lysine-rich nuclear localization signal (NLS) sequence between Lys³⁷⁴ and Lys³⁸¹ in the X-Y linker region. Truncation of EF1 resulted in the loss of NTA, and point mutation revealed a responsible sequence in the N terminus of EF1. However, even if EF1 was present, NTA was lost when EF2-4 or C2 domain was deleted. Both NTA and Ca²⁺ oscillation-inducing ability are lost in these truncation or deletion mutants. Similar results were obtained in cultured COS cells after transfection with cDNA of mutants. It is predicted from the 3-D structure of PLCδ1 that PLCζ is folded at the hinge region in the X-Y linker and that EF-hand domains and C2 domain make extensive contact. Besides NLS, highly coordinated overall structure of PLCζ is responsible for NTA as well as Ca²⁺ oscillation-inducing activity. [J Physiol Sci. 2006;56 Suppl:S116]