Fig 2: Model for LINKIN-dependent cell adhesion during cell migration.We hypothesize that LNKN-1 promotes cell adhesion through the interactions of its extracellular domain with other cells or extracellular matrix and through its intracellular domain by interacting with the microtubule cytoskeleton, which connects it to the chromosome. This allows the use of the nucleus as a huge anchor that can withstand the force of collective cell migration. RUVB-1/2 interact with LNKN-1 at the plasm membrane, and the microtubule cytoskeleton interacts with the chromosome potentially through the anaphase-promoting complex. NDK-1 and TAT-5 also promote cell adhesion and are likely associated with LNKN-1 at the plasma membrane, while condensin and cohesin likely promote cell adhesion through their interaction with chromosomes.

Fig 3: Potential LNKN-1 interactors displayed a similar gonad migration phenotype in hermaphrodites.(A–B). Loss of lnkn-1 resulted in premature stopping of gonad migration in hermaphrodites (A) lnkn-1(+) control. (B) lnkn-1(sy1596) worms have a shortened dorsal-distal half of the gonad. (C-D) The premature stopping phenotype is defined by comparing the value of Centripetal migration (%) of the mutants to that of the wild-type (N2). The Centripetal migration (%) is determined by the ratio of dorsal-distal gonad arm length to that of ventral-proximal gonad arm length. The arm lengths were approximated as the distance from the vulval to the ventral-dorsal turn of the U-shaped gonad for the ventral-proximal gonad arm length; and the distance from the distal end of the gonad to the ventral-dorsal turn for the dorsal-distal gonad arm length. See materials and methods for detail. (D) Examination of gonad migration in hermaphrodites. Mutation in some of the candidate genes also causes pathfinding or other gonadal development defects in a number of animals. For the purpose of this examination, only gonads that made the centripetal migration was included. The pathfinding effects were consistently observed in cdc-25.1(ok1888) and rod-1(tm6186) and occurred at a lower frequency in tat-5(tm1741) and apc-2(ok1657) mutant animals. Rod-1(tm6186) animals were also observed to have missing gonad arms (n≥10. All comparisons were with the wild-type, **: p<0.01, ***: p<0.001, ****: p<0.0001, Student’s t-test).

Fig 4: Identifying potential ITFG1 interactors with mass spectrometry.(A–B) fold change of the protein level in log2 between ITFG1-Myc-expressing cells over non-expressing control MDA-MB-231 cells. Each dot represents one protein. Red-ITFG1, Purple- RUVBL1, Green-RUVBL2. FC, fold change. (A) Transient ITFG1-Myc-expressing cells (B) Stable ITFG1-Myc-expressing cells. (C) Reactome pathway and GO biological process enrichment of the overlapping 180 proteins in ITFG-Myc expressing MDA-MB-231 cells. Data present the top 20 statistically enriched terms (P-adjusted value < 0.05).

Fig 5: Mutations in the orthologs of ITFG1 interactors resulted in phenotypes similar to that of lnkn-1 mutants.(A) Homozygotic animals of lethal/ sterile mutations used in this study descended from balanced heterozygotic mothers. Homozygotic animals were identified through the lack of fluorescently labeled balancers (green in the cartoon). In the maternal effect experiments, the homozygotic hermaphrodites were mated with wild-type males carrying a fluorescent genetic marker (oxTi719 [eft-3p::tdTomato::H2B]) (Frokjaer-Jensen et al., 2014). (B) LNKN-1 is required maternally. Heterozygotic cross-progenies of lnkn-1(sy1596) hermaphrodites and lnkn-1(+/+) males fail to develop and die without hatching (The embryos carrying the orange-colored male-derived oxTi719 marker can be seen). (C) Phenotypical analysis of the orthologs of ITFG1 interactors in C. elegans. Phenotype 2 and 5 was defined by having F1 progenies that hatch; Phenotype 3 was defined by embryos/ embryo-like objects or oocytes observed outside of the unmated hermaphrodite. Embryos and, in particular, oocytes may not be completely absent in all animals. This was especially the case in unc-94(ok1210). Some rod-1(tm6186) animals also produce oocytes and embryo-like objects. Phenotype 4 was defined by the observation of F1 embryos/animals carrying the male-derived oxTi719 marker. *: Very rarely; #: maternal effect sterile.