Interaction between Discs large and Pins/LGN/GPSM2: a comparison across species

Abstract

The orientation of the mitotic spindle determines the direction of cell division, and therefore contributes to tissue shape and cell fate. Interaction between the multifunctional scaffolding protein Discs large (Dlg) and the canonical spindle orienting factor GPSM2 (called Pins in Drosophila and LGN in vertebrates) has been established in bilaterian models, but its function remains unclear. We used a phylogenetic approach to test whether the interaction is obligate in animals, and in particular whether Pins/LGN/GPSM2 evolved in multicellular organisms as a Dlg-binding protein. We show that Dlg diverged in C. elegans and the syncytial sponge O. minuta and propose that this divergence may correspond to differences in spindle orientation requirements between these organisms and the canonical pathways described in bilaterians. We also demonstrate that Pins/LGN/GPSM2 is present in basal animals, but the established Dlg-interaction site cannot be found in either Placozoa or Porifera. Our results suggest that the interaction between Pins/LGN/GPSM2 and Dlg appeared in Cnidaria, and we therefore speculate that it may have evolved to promote accurate division orientation in the nervous system. This work reveals the evolutionary history of the Pins/LGN/GPSM2-Dlg interaction and suggests new possibilities for its importance in spindle orientation during epithelial and neural tissue development.

AGS3 antagonizes LGN to balance oriented cell divisions and cell fate choices in mammalian epidermis

“…The spindle orienting function of Gpsm2 and its homologs is highly conserved throughout evolution, and may have evolved as early as the time when bilateria and cnidaria diverged (Schiller & Bergstralh, 2021; Wavreil & Yajima, 2020). While LGN and its orthologs have an evolutionarily-conserved role in oriented cell divisions, the manner and pattern in which LGN localizes to the cell cortex varies among epithelial tissues.…”

“…In addition to Insc and Gαi family proteins, a growing list of proteins— including Dlg, E-cadherin, Frmpd1/4 and afadin—have been found to promote the cortical localization of LGN in different contexts (Carminati et al, 2016; Gloerich, Bianchini, Siemers, Cohen, & Nelson, 2017; Schiller & Bergstralh, 2021; Wee, Johnston, Prehoda, & Doe, 2011; Yuzawa et al, 2011). Although autoinhibition and phosphorylation are mechanisms known to regulate LGN’s ability to interact with proteins that promote its membrane association (Bergstralh et al, 2017; Johnston, Hirono, Prehoda, & Doe, 2009; Pan et al, 2013), comparatively less is known about negative regulators.…”

“…While it is not known whether aPKC phosphorylation of LGN impacts its spindle orienting capacity, loss of Prkci in the epidermis does result in a spindle orientation phenotype (Niessen et al, 2013). On the other hand, the Dlg-LGN interaction is highly conserved throughout evolution (Schiller & Bergstralh, 2021), and a non-phosphorylatable pinsS436A mutant is non-functional in a Drosophila S2 induced-polarity assay, while the equivalent mutation in vertebrate LGN (S401A) induces spindle misorientation in MDCK cells and the chick neural tube (Hao et al, 2010; Saadaoui et al, 2014). This serine residue is conserved in the AGS3 linker, but there are some differences in the flanking residues, and it has been reported that the AGS3 linker binds Dlg with ∼500-fold lower efficiency than LGN (Zhu, Shang, et al, 2011).…”

“…LGN is a member of the AGS (Activator of G-protein signaling) family of proteins, named because they promote G-protein signaling in a receptor-independent manner. In vertebrates, LGN has a closely-related paralog, AGS3 (Gpsm1), and the two proteins share high protein homology and a conserved domain structure, consisting of seven to eight N-terminal tetra-tricopeptide repeats (TPR) and four C-terminal GoLoco motifs—also known as the G-protein regulatory (GPR) region—separated by a flexible linker (Blumer, Cismowski, Sato, & Lanier, 2005; Schiller & Bergstralh, 2021; Wavreil & Yajima, 2020). Biochemical and structural studies predict that AGS3 retains the ability to interact with many of the same binding partners as LGN, including Insc and NuMA (Adhikari & Sprang, 2003; Culurgioni, Alfieri, Pendolino, Laddomada, & Mapelli, 2011; Izaki, Kamakura, Kohjima, & Sumimoto, 2006; Saadaoui et al, 2017; Yuzawa, Kamakura, Iwakiri, Hayase, & Sumimoto, 2011; Zhu, Wen, et al, 2011).…”

The spindle orienting machinery requires activation, not just localization

“…The finding that there are two distinct mechanisms for activating pulling raises the question of how and for what purpose they evolved. Phylogenetic analysis indicates that the interaction between Pins and Dlg is evolutionarily ancient; it is predicted to have evolved in Cnidaria, predating the appearance of Inscuteable orthologs (Schiller and Bergstralh, 2021). It is therefore tempting to speculate that Inscuteable-mediated spindle orientation evolved to promote cell type diversity, particularly in the more complex bilaterian nervous system.…”