Mardick, J.I. et al. (2019) International Worm Meeting "The RalGEF-Ral signaling module confers positional information in cell migration."

Wightman, B., Mardick, J.I., Reiner, D.J.

[International Worm Meeting, 2019]

The Ras small GTPase is the most mutated oncoprotein, driving both tumorigenesis and metastasis. Oncogenic Ras effectors Raf and PI3 Kinase cascades are well studied and pharmacologically targeted. Ras directly binds a third oncogenic effector, RalGEF, which in turn activates a "Ras-Like" cousin, Ral. Yet the downstream consequences of RalGEF-Ral activation are poorly understood. Consequently, we are studying the functions of RalGEF-Ral in C. elegans development. We previously reported that the LET-60/Ras-RGL-1/RalGEF-RAL-1/Ral signal promotes 2? fate in vulval cell fate patterning. MIG-2 is an ortholog of mammalian RhoG, a known regulator of actin dynamics and cell migration. The gm38gf activating mutation in MIG-2 confers modest defects in migration of canal associated neurons (CANs), a bilaterally symmetric pair of neurons that migrate posteriorly from the head to the middle of the animal along the antero-posterior axis. We observed that a deletion of nine genes on the X chromosome, including rgl-1, enhanced the migration defect of mig-2(gm38gf), as did rgl-1 deletions ok1921 and tm2255. Deletions in rgl-1 similarly enhanced CAN mutation defects caused by mutation of VAB-8/kinesin-like and UNC-34/Enabled. Yet in a wild-type background, deletions of rgl-1 did not confer cell migration defects. Thus, RGL-1 is a modifier but not a central player in cell migration. A GEF-specific mutation in RGL-1 and a reduced function mutation in RAL-1 acted similarly, confirming that canonical activation of RAL-1 by RGL-1 is required for optimal cell migration. A published gain-of-function mutation in RAL-1 conferred the same defect as loss of RGL-1 and RAL-1 function, arguing that the role of Ral in CAN migration is instructive, not permissive. Our future goals are to test whether Ras itself regulates CAN cell migration, determine whether RGL-1-RAL-1 functions cell autonomously in the CANs, and extend our analysis to additional cell and axonal migration events. Mirroring its established role in metastasis, we find that RalGEF-Ral regulates cell migration during normal animal development.

Mardick JI et al. (2021) Dev Biol "Parallel Rap1>RalGEF>Ral and Ras signals sculpt the C. elegans nervous system."

Wightman B, Mardick JI, Reiner DJ, Rasmussen NR

[Dev Biol, 2021]

Ras is the most commonly mutated oncogene in humans and uses three oncogenic effectors: Raf, PI3K, and RalGEF activation of Ral. Understanding the importance of RalGEF>Ral signaling in cancer is hampered by the paucity of knowledge about their function in animal development, particularly in cell movements. We found that mutations that disrupt function of RalGEF or Ral enhance migration phenotypes of mutants for genes with established roles in cell migration. We used as a model the migration of the canal associated neurons (CANs), and validated our results in HSN cell migration, neurite guidance, and general animal locomotion. These functions of RalGEF and Ral are specific to their control of Ral signaling output rather than other published functions of these proteins. In this capacity Ral functions cell autonomously as a permissive developmental signal. In contrast, we observed Ras, the canonical activator of RalGEF>Ral signaling in cancer, to function as an instructive signal. Furthermore, we unexpectedly identified a function for the close Ras relative, Rap1, consistent with activation of RalGEF>Ral. These studies define functions of RalGEF>Ral, Rap1 and Ras signaling in morphogenetic processes that fashion the nervous system. We have also defined a model for studying how small GTPases partner with downstream effectors. Taken together, this analysis defines novel molecules and relationships in signaling networks that control cell movements during development of the nervous system.