De novo genesis of retinal ganglion cells by targeted expression of Klf4 in vivo

Abstract

Retinal ganglion cell (RGC) degeneration is a hallmark of glaucoma, the most prevalent cause of irreversible blindness. Thus, therapeutic strategies are needed to protect and replace these projection neurons. One innovative approach is to promote de novo genesis of RGCs via manipulation of endogenous cell sources. Here, we demonstrate that the pluripotency regulator gene Krüppel-like factor 4 (Klf4) is sufficient to change the potency of lineage-restricted retinal progenitor cells to generate RGCs in vivo. Transcriptome analysis disclosed that the overexpression of Klf4 induces crucial regulators of RGC competence and specification, including Atoh7 and Eya2. In contrast, loss-offunction studies in mice and zebrafish demonstrated that Klf4 is not essential for generation or differentiation of RGCs during retinogenesis. Nevertheless, induced RGCs (iRGCs) generated upon Klf4 overexpression migrate to the proper layer and project axons aligned with endogenous fascicles that reach the optic nerve head. Notably, iRGCs survive for up to 30 days after in vivo generation. We identified Klf4 as a promising candidate for reprogramming retinal cells and regenerating RGCs in the retina. This article has an associated 'The people behind the papers' interview.

On the Generation and Regeneration of Retinal Ganglion Cells

“…This study showed that Klf4 induced the reactivation of the early neurogenic program in late progenitors, changing their competence to generate RGCs that properly localized to the inner retina and projected axons into the optic nerve head (Rocha-Martins et al, 2019). The precise mechanism underlying the effect of Klf4 in late progenitors is still unknown, but we hypothesize that Klf4 reactivates the molecular program for RGC differentiation through its properties as a pioneer factor, combined with the direct or indirect induction of Atoh7 (Chronis et al, 2017;Rocha-Martins et al, 2019). Although these results are promising, the detailed characterization of the transcriptional signature, subtype, and function of these induced RGCs, as well as their capacity to connect within the retina and with their brain targets remains to be defined.…”

“…This may be due to redundancy with other Klf family members (Jiang et al, 2008), since many are expressed in the developing retina (Moore et al, 2009;Njaine et al, 2014). We recently showed that overexpression of Klf4 in late retinal progenitors generates induced RGCs outside of their developmental window (Figure 1; Rocha-Martins et al, 2019). This study showed that Klf4 induced the reactivation of the early neurogenic program in late progenitors, changing their competence to generate RGCs that properly localized to the inner retina and projected axons into the optic nerve head (Rocha-Martins et al, 2019).…”

“…The potential roles of other elements of this network, like fly Krüppel and Pdm, remain unknown. Recently, Klf4, a member of the family of Krüppel-like factors was studied in the mouse retina, but no critical function in cell fate determination was described (Moore et al, 2009;Fang et al, 2016;Rocha-Martins et al, 2019). This may be due to redundancy with other Klf family members (Jiang et al, 2008), since many are expressed in the developing retina (Moore et al, 2009;Njaine et al, 2014).…”

“…We recently showed that overexpression of Klf4 in late retinal progenitors generates induced RGCs outside of their developmental window (Figure 1; Rocha-Martins et al, 2019). This study showed that Klf4 induced the reactivation of the early neurogenic program in late progenitors, changing their competence to generate RGCs that properly localized to the inner retina and projected axons into the optic nerve head (Rocha-Martins et al, 2019). The precise mechanism underlying the effect of Klf4 in late progenitors is still unknown, but we hypothesize that Klf4 reactivates the molecular program for RGC differentiation through its properties as a pioneer factor, combined with the direct or indirect induction of Atoh7 (Chronis et al, 2017;Rocha-Martins et al, 2019).…”

RINT1 Loss Impairs Retinogenesis Through TRP53-Mediated Apoptosis

“…Measurements of visual acuity by optomotor response were performed using OptoMotry as previously described (Cavalheiro et al, 2017;Rocha-Martins et al, 2019). Visual accuracy threshold was determined by systematic increments of the spatial frequency until the animal no longer responded.…”