Isolation, characterization and molecular cloning of cathepsin D from lizard ovary: Changes in enzyme activity and mRNA expression throughout ovarian cycle

Abstract

During vitellogenesis, the oocytes of oviparous species accumulate in the cytoplasm a large amount of proteic nutrients synthetized in the liver. Once incorporated into the oocytes, these nutrients, especially represented by vitellogenin (VTG) and very low-density lipoprotein (VLDL), are cleaved into a characteristic set of polypeptides forming yolk platelets. We have studied the molecular mechanisms involved in yolk formation in a reptilian species Podarcis sicula, a lizard characterized by a seasonal reproductive cycle. Our results demonstrate the existence in the lizard ovary of an aspartic proteinase having a maximal activity at acidic pH and a molecular mass of 40 kDa. The full-length aspartic proteinase cDNA produced from total RNA by RT-PCR is 1,442 base pairs long and encodes a protein of 403 amino acids. A comparison of the proteic sequence with aspartic proteinases from various sources demonstrates that the lizard enzyme is a cathepsin D. Lizard ovarian cathepsin D activity is maximal in June, in coincidence with vitellogenesis and ovulation, and is especially abundant in vitellogenic follicles and in eggs. Ovarian cathepsin D activity can be enhanced during the resting period by treatment with FSH in vivo. Northern blot analysis shows that cathepsin D mRNA is exceedingly abundant during the reproductive period, and accumulates preferentially in previtellogenic oocytes.

Evolutionary fate of duplicate genes encoding aspartic proteinases. Nothepsin case study

“…The results reported here show the presence in lizard of an aspartic proteinase form distinct from the cathepsin D previously described in the same organism (De Stasio et al, 1999); sequence analysis established for the new lacertilian sequence features typical of nothepsin. However, at variance with fish, lizard nothepsin is constitutively expressed in both males and females.…”

Stable isotope tracers reveal a trade‐off between reproduction and immunity in a reptile with competing needs

“…Both reproduction and self-maintenance require protein, and the immunosuppressive effects of pregnancy are well documented Martin, Weil, & Nelson, 2008), although most studies simply compare pregnant females with non-pregnant ones rather than assessing how allocation of energy to developing offspring change throughout pregnancy. Little is known about the timing of protein allocation to egg yolk in reptiles, and our assumption of a constant allocation rate is almost certainly incorrect (De Stasio, Borrelli, Kille, Parisi, & Filosa, 1999;Lourdais, Bonnet, Shine, & Taylor, 2003;Van Dyke & Beaupre, 2011). Vitellogenesis is the most resource-demanding stage of reproduction for many organisms, including reptiles (Thompson et al, 2001;Van Dyke & Beaupre, 2011), and there is some evidence that early reproduction is more energetically costly than later stages in small, heterothermic mammals (Christe, Arlettaz, & Vogel, 2000;Racey & Speakman, 1987).…”

Thyroglobulin type‐1 domain protease inhibitors exhibit specific expression in the cortical ooplasm of vitellogenic rainbow trout oocytes

“…Multiple studies have identified cathepsin proteases as key mediators of yolk processing during vitellogenic oocyte development in a wide variety of species (Fagotto, 1990;Nakamura et al, 1996;De Stasio et al, 1999;Gerhartz et al, 1999;Carnevali et al, 1999b). What remains enigmatic are the mechanisms by which multiple cathepsin proteases are differentially controlled during key developmental events involving proteolysis.…”

“…There is considerable evidence to suggest that the proteolytic processing of both vitellogenin and yolk platelets is mediated by cathepsins, a diverse family of serine, aspartate, and cysteine proteases. Cathepsins have been implicated in yolk processing in fishes (Kestemont et al, 1999;Carnevali et al, 1999a,b;Kwon et al, 2001;Hiramatsu et al, 2002;Wood and Van Der Kraak, 2003), amphibians (Yoshizaki and Yonezawa, 1996), reptiles (De Stasio et al, 1999), birds (Gerhartz et al, 1999), and insects (Yamamoto et al, 1994;Ribolla et al, 2001), suggesting an evolutionarily conserved role for these enzymes in vitellogenin and yolk processing.…”