br Learning from our ancestors

Learning from our ancestors As mentioned earlier, homologues of all γ-secretase components have been identified in plants and protozoans, some of which have emerged as model systems for the study of γ-secretase independent functions of the presenilins [27]. The moss P. patens was the first plant species in which conservation of γ-secretase complex components and a γ-secretase independent function for the presenilin homologue (PpPS) and nicastrin was demonstrated [41]. A more recent study in A. thaliana similarly showed the conservation of γ-secretase complex components in another plant species [42], [43]. Importantly, certain highly conserved amino MI-77301 motifs crucial for γ-secretase proteolytic activity, substrate recognition and complex assembly in mammals are similarly conserved in plant species [68]. However, phylogenetic analysis of presenilins indicates that plant and animal homologues fall into two divergent clades [43]. Interestingly, P. patens genome contains only one copy of presenilin (PpPS) and does not possess homologues of several mammalian γ-secretase substrates including APP and Notch [41]. In reconstitution studies PpPS was defective in cleavage of a Notch1-based substrate in PS-deficient mouse embryonic fibroblasts (MEFs), but was able to restore normal proliferation rates in PS-deficient MEFs, a function thought to be independent of γ-secretase proteolytic activity [41]. In the phenotypic characterization of a null mutant of presenilin (Ppps), P. patens displayed abnormal growth pattern, impaired chloroplast movement and decreased endocytosis [41]. In this regard, in Arabidopsis studies employing exogenous expression of γ-secretase subunits impairment of vacuole trafficking was observed [43]. Together this data from two plant species intimates that the predominant function of presenilin in plants is not related to γ-secretase protease activity but perhaps points to an ancestral role for presenilins and the γ-secretase complex in protein endocytosis and trafficking. Consistent with this hypothesis, γ-secretase is present and active in the slime mould D. discoideum, [44] where the presenilins have been shown to be essential for Dictyostelium cell fate determination and the regulation of phagocytosis. However, in contrast to plants, Dictyostelium γ-secretase was still proteolytically active against animal substrates [44]. Again, this suggests an ancestral role for presenilins that extends well beyond what is known about γ-secretase protease activity from predominantly animal studies. Nevertheless, from animal studies there is also a growing body of evidence showing that the presenilins and γ-secretase may be involved in the regulation of endocytosis, protein trafficking and degradation [69], [70], [71], leading to the proposal that signalling functions associated with γ-secretase proteolytic activity and substrate cleavage arose later in evolution [26], [44]. In keeping with this, while there are over 90 reported γ-secretase substrates no biological function has been attributed to the majority of these γ-secretase-generated ICDs [10], [22]. However, many substrate-derived ICDs are rapidly degraded and support the proposal that the predominant function attributed to γ-secretase would be to facilitate the removal and degradation of membrane bound protein fragments subsequent to ectodomain shedding, thereby acting as a ‘membrane proteasome’ [72]. Furthermore, mice deficient in Notch signalling or deficient in either of the γ-secretase components, Aph-1, Pen-2 or Nicastrin, are able to generate anterior somites, whereas Psen1 and Psen2 double-deficient animals are not, suggesting that the presenilins contribute to the generation of anterior somites, independent of their role in γ-secretase [73]. In summary, from an evolutionary perspective, data supports the proposal that presenilins have an important regulatory role in protein endocytosis, trafficking and degradation, which preceded signalling functions associated with γ-secretase substrate cleavage and proteolytic activities.