RASGRF2 controls nuclear migration in postnatal retinal cone photoreceptors

Abstract

Detailed immunocytochemical analyses comparing wild-type (WT), GRF1-knockout (KO), GRF2-KO and GRF1/2 double-knockout (DKO) mouse retinas uncovered the specific accumulation of misplaced, 'ectopic' cone photoreceptor nuclei in the photoreceptor segment (PS) area of retinas from GRF2-KO and GRF1/2-DKO, but not of WT or GRF1-KO mice. Localization of ectopic nuclei in the PS area of GRF2-depleted retinas occurred postnatally and peaked between postnatal day (P)11 and P15. Mechanistically, the generation of this phenotype involved disruption of the outer limiting membrane and intrusion into the PS layer by cone nuclei displaying significant perinuclear accumulation of signaling molecules known to participate in nuclear migration and cytoskeletal reorganization, such as PAR3, PAR6 and activated, phosphorylated forms of PAK, MLC2 and VASP. Electroretinographic recordings showed specific impairment of cone-mediated retinal function in GRF2-KO and GRF1/2-DKO retinas compared with WT controls. These data identify defective cone nuclear migration as a novel phenotype in mouse retinas lacking GRF2 and support a crucial role of GRF2 in control of the nuclear migration processes required for proper postnatal development and function of retinal cone photoreceptors.

Generation of a Cone Photoreceptor-specific GNGT2 Reporter Line in Human Pluripotent Stem Cells

“…While postmitotic movements of cone nuclei have been reported in mice, the signals and molecular pathways controlling this process have not been resolved [38,39]. A study of ectopically located cones in GRF2-KO mice suggested a potential role for GRF2 and CDC42 in the formation/structure of PARcontaining, polarity-related macromolecular complexes [40,41], and the serine/threonine kinase LKB1 as well as one of its substrates, AMPK, have also been identified as regulators of cone nuclear positioning [42]. Our observations suggest that in human retinae apical cones extend processes basally, prior to relocation, in an active process rather than one based on simple somal displacement by newly generated rods.…”

Rap Guanine Nucleotide Exchange Factor 3 (Rapgef3)

“…Thus, RasGrf2 KO mice show lower levels of ERK activation upon NMDA-induction and defective LTP in the CA1 region of the hippocampus (Feig 2011). Additionally, RasGrf2 has also been shown to be important for development of LTP and survival of newborn neurons in the dentate girus (Darcy et al 2014b), for morphogenesis of the retina (Jimeno et al 2016) and for the control of dopaminergic and noradrenergic responses to alcohol (Stacey et al 2012).…”

Disrupting the ciliary gradient of active Arl3 affects rod photoreceptor nuclear migration

“…During development, all retinal neurons are born apically and then migrate to their final retinal layer where they elaborate and make connections (Baye and Link, 2008). For cones, whose nuclei are arranged at the apical side of the ONL, it is known that their nuclear position can be influenced by either knocking out RASGRF2 the GEF for the Ras/Rho/Rac family of small GTPases (Jimeno et al, 2016), altering dopamine signaling through the D4 receptor (Tufford et al, 2018), or uncoupling the nucleus from microtubules (Yu et al, 2011; Razafsky et al, 2012). Less is known about how rod nuclear position is determined, but microtubule coupling to the nucleus is important.…”