Studies on the regulation of E3 ubiquitin ligase APC3 and its interacting proteins on the tetraspore formation and release in Gracilariopsis lemaneiformis (Rhodophyta).

E3 ubiquitin ligases play significant roles in development of high plants and animals. We recently found that E3 ubiquitin ligase APC3, the subunit of the anaphase promoting complex/cyclosome, was involved in tetraspore formation and release in Gracilariopsis lemaneiformis, an economically important red alga. GlAPC3 showed opposite expression pattern in low-fertility cultivar 981 and high-fertility strain WLP during the process of tetraspore formation and release, up-regulated in 981 and down-regulated in WLP. Five proteins related to chromosome segregation, SMC3, NUF2, APC2, APC8 and APC10, were detected to interact with APC3, which were all located in the nucleus. NUF2 and CDC20 were the substrates of APC3, combined with Lysine-11, Lysine-48 and Lysine-63 of ubiquitin chains containing two or four ubiquitin. The key amino acids for ubiquitination of APC3 covered 474th Aspartate, 502nd tyrosine and 506th leucine, any mutation of which resulted in a loss of ubiquitination. During the process of tetraspore formation and release, SMC3 was significantly up-regulated only in 981, low number of tetraspore release. NUF2 and APC2 were significantly down-regulated only in WLP, with high frequency and large amount of tetraspores release. The data provided that APC3, SMC3 and NUF2 might be the key gene affecting the fertility of Gp. lemaneiformis. The study helps to explore the regulation mechanism of APC3 with SMC3 and NUF2 by the process of chromatids segregation in regulating tetraspore formation and release of Gp. lemaneiformis.