OTX2 non-cell autonomous activity regulates inner retinal function

Abstract

a These authors contributed equally to this work. Abstract: 179 words; significance statement: 118 words; introduction: 463 words; discussion: 1344 words.Acknowledgements: none.Conflict of interest: KLM and AP are listed on patents for the use of homeoproteins to treat neurodegenerative disease and each holds equity in a start up with that aim. AbstractOTX2 is a homeoprotein transcription factor expressed in photoreceptors and bipolar cells in the retina. OTX2, like many other homeoproteins, transfers between cells and exerts non-cell autonomous effects such as promoting survival of retinal ganglion cells that do not express the protein. Here we used a genetic approach to target extracellular OTX2 in the retina by conditional expression of a secreted single chain anti-OTX2 antibody. Compared to control mice, the expression of this antibody by Parvalbumin-expressing neurons in the retina is followed by a reduction in visual acuity in one-month-old mice with no alteration of the retinal structure or cell type number or aspect. A-and b-waves measured by electroretinogram were also indistinguishable from control mice, suggesting no functional deficit of photoreceptors and bipolar cells. Mice expressing the OTX2-neutralizing antibody did show a significant doubling in the flicker amplitude, consistent with a change in inner retinal function. Our results demonstrate that interfering in vivo with OTX2 non-cell autonomous activity in the postnatal retina leads to an alteration in inner retinal cell functions and causes a deficit in visual acuity. OTX2 non-cell autonomous activity regulates inner retinal function 4 Significance statement OTX2 is a homeoprotein transcription factor expressed in retinal photoreceptors and bipolar cells. Although the Otx2 locus is silent in the inner retina, the protein is detected in cells of the ganglion cell layer consistent with the ability of this class of proteins to transfer between cells. We expressed a secreted single chain antibody (scFv) against OTX2 in the retina to neutralize extracellular OTX2. Antibody expression leads to reduced visual acuity with no change in retinal structure, or photoreceptor or bipolar physiology; however, activity in the inner retina was altered. Thus, interfering with OTX2 non-cell autonomous activity in postnatal retina alters inner retinal function and causes vision loss, highlighting the physiological value of homeoprotein direct non-cell autonomous signaling. OTX2 non-cell autonomous activity regulates inner retinal function

Homeoprotein neuroprotection of embryonic neuronal cells

“…Exogenous ENGRAILED protects DA neurons in vitro against MPP+ and rotenone and in vivo against 6-OHDA, MPTP, A30P α-synuclein and progressive degeneration associated with the loss of one EN1 allele (Sonnier et al, 2007;Alvarez-Fischer et al, 2011;Rekaik et al, 2015, Thomasson et al, 2019. OTX2 promotes the survival of adult dissociated RGCs in vitro, protects RGC in vivo against NMDA excitotoxicity and mDA neurons against 6-OHDA (Torero-Ibad et al, 2011;Rekaik et al, 2015). This similar pro-survival activity of two distinct transcription factors of different HP families led us to develop an in vitro assay to assess the ability of several HPs belonging to different classes to protect embryonic neurons against cell death and DNA damage caused by H 2 O 2 oxidative stress.…”

“…Mouse EN1, hEN1 and mouse or chEN2 are neuroprotective towards midbrain dopaminergic cells in vitro and in vivo (Sonnier et al, 2007;Alvarez-Fischer et al, 2011;Rekaik et al, 2015). Protection was also observed for OTX2 on DA midbrain cells in vivo and on RGCs in vitro and in vivo (Sonnier et al, 2007;Alvarez-Fischer et al, 2011;Torero et al, 2011;Rekaik et al, 2015). This raised the possibility that protective activity may be a property shared among a number of HPs.…”

“…Chicken ENGRAILED2 (chEN2) and mutant chicken ENGRAILED2 (SR-EN2), mouse EN1 (mEN1), human EN1 (hEN1) and mouse OTX2 (mOTX2) were prepared as described (Joliot et al, 1998;Torero et al, 2011). Cell-permeable recombinant human c-MYC was purchased from Abcam (ab169901) and human GBX2 (hGBX2) and LHX9 (hLHX9) were purchased from Proteogenix.…”

“…(EN1) and ENGRAILED2 (EN2) (collectively ENGRAILED), as therapeutic proteins in animal models of Parkinson disease (ENGRAILED) and glaucoma (OTX2) (Sonnier et al, 2007;Alvarez-Fischer et al, 2011;Torero-Ibad et al, 2011;Thomasson et al, 2019).…”

“…Because OTX2 has a similar survival effect on mDA neurons and retinal ganglion cells (Rekaik et al, 2015;Torero-Ibad et al, 2011), it could be that protection against oxidative stress is a shared property of several HPs with little HP and/or neuronal specificity. To test this hypothesis, the protective effect of EN1, EN2, OTX2, GBX2 and LHX9 was evaluated on 4 midbrain and striatal neurons in culture as well as testing hEN1 on a neuronal cell line.…”