Assembly of the TgrB1–TgrC1 cell adhesion complex during Dictyostelium discoideum development

Abstract

In Dictyostelium discoideum, TgrB1 and TgrC1 are partners of a heterophilic cell-adhesion system. To investigate its assembly process, the split GFP complementation assay was used to track the oligomeric status of both proteins. The ability of TgrC1 to form cis-homodimers spontaneously was demonstrated by fluorescence complementation studies and confirmed by chemical cross-linking. In contrast, TgrB1 failed to form cis-homodimers in the absence of TgrC1. Treatment of cell aggregates with antibodies against TgrB1 or TgrC1 did not affect TgrC1 dimerization, but inhibited TgrB1 dimer formation, suggesting that TgrB1 cis-homodimerization is dependent on trans-interaction with TgrC1. When TgrB1 and TgrC1 conjugated with the complementary halves of GFP were co-expressed in cells, cis-heterodimers were not detected. However, weak FRET signals were detected in cells expressing TgrB1-RFP and TgrC1-GFP, suggesting that TgrB1 dimers and TgrC1 dimers were arranged juxtapose to each other in the adhesion complex. The results of the present study suggest that the assembly process is initiated upon trans-interaction of monomeric TgrB1 with TgrC1 homodimers on adjacent cells, which triggers the formation of TgrB1 dimers. The homodimerization of TgrB1 in turn induces the clustering of TgrB1 and TgrC1, and the coalescence of TgrB1-TgrC1 clusters results in the formation of large adhesion complexes.

Allorecognition, via TgrB1 and TgrC1, mediates the transition from unicellularity to multicellularity in the social amoebae Dictyostelium discoideum

“…This is consistent with the report that TgrB1 and TgrC1 establish associations within the membrane of one cell that is stimulated by their interactions between cells, suggesting the formation of TgrB1/TgrC1 adhesion patches (Chen et al, 2014).…”

“…The allorecognition proteins TgrB1 and TgrC1 are polymorphic in natural populations and mediate heterotypic cell-cell adhesion (Wang et al, 2000;Benabentos et al, 2009;Chen et al, 2013Chen et al, , 2014. Thus, the TgrB1 and TgrC1 proteins provide a possible mechanism for the cooperation that has been described in this system (Buss, 1982;Mehdiabadi et al, 2006;Gilbert et al, 2007;Ostrowski et al, 2008).…”

Tissue self-organization based on collective cell migration by contact activation of locomotion and chemotaxis

“…TgrB1-RFP TgrC1-GFP expression vectors were constructed as follows. The tgrB1 gene including the promoter and the ORF except the stop codon was amplified with an additional N-terminal Nhe I site and a C-terminal (G 4 S) 2 linker sequence[52] and a Cla I site by PCR. Vector pHygTm(+) was cut by Xba I/ Cla I to replace the act15 promoter with the amplified tgrB1 fragment to obtain pHyg-[tgrB1p]:tgrB1-(G 4 S) 2 -mRFP1.…”

Altered N-glycosylation modulates TgrB1- and TgrC1-mediated development but not allorecognition in Dictyostelium

“…The TgrB1 and TgrC1 cell adhesion system of Dictyostelium is an essential component of development, with roles in morphogenesis and allorecognition (Dynes et al, 1994;Hirose et al, 2011). Although the precise signaling pathways that mediate these roles are uncertain, it is clear that allorecognition is mediated by allele-specific interactions between TgrB1 molecules on the outer membrane of one cell and matching TgrC1 molecules on the outer membrane of a neighboring cell (Chen et al, 2013(Chen et al, , 2014Hirose et al, 2011). It was therefore surprising to find that allorecognition, which is mediated by TgrB1 and TgrC1, was not affected by compromised N-linked glycosylation.…”

“…The tight mounds subsequently elongate to form migrating slugs and culminate into fruiting bodies consisting of spore-containing sori held aloft by cellular stalks. The tgrB1 and tgrC1 genes encode two immunoglobulin-like cell adhesion molecules (Ig-like CAMs) that form a heterophilic cell-cell adhesion complex (Chen et al, 2013(Chen et al, , 2014 and are crucial for the initiation of cell differentiation. Cells with disrupted tgrB1 or tgrC1 genes fail to express cell-type-specific genes and are unable to progress beyond the loose mound stage (Dynes et al, 1994;Iranfar et al, 2006).…”