Hexokinase 2 is dispensable for photoreceptor development but is required for survival during aging and outer retinal stress

Abstract

Photoreceptor death is the ultimate cause of vision loss in many retinal degenerative conditions. Identifying novel therapeutic avenues for prolonging photoreceptor health and function has the potential to improve vision and quality of life for patients suffering from degenerative retinal disorders. Photoreceptors are metabolically unique among other neurons in that they process the majority of their glucose via aerobic glycolysis. One of the main regulators of aerobic glycolysis is hexokinase 2 (HK2). Beyond its enzymatic function of phosphorylating glucose to glucose-6-phosphate, HK2 has additional non-enzymatic roles, including the regulation of apoptotic signaling via AKT signaling. Determining the role of HK2 in photoreceptor homeostasis may identify novel signaling pathways that can be targeted with neuroprotective agents to boost photoreceptor survival during metabolic stress. Here we show that following experimental retinal detachment, p-AKT is upregulated and HK2 translocates to mitochondria. Inhibition of AKT phosphorylation in 661W photoreceptor-like cells results in translocation of mitochondrial HK2 to the cytoplasm, increased caspase activity, and decreased cell viability. Rod-photoreceptors lacking HK2 upregulate HK1 and appear to develop normally. Interestingly, we found that HK2-deficient photoreceptors are more susceptible to acute nutrient deprivation in the experimental retinal detachment model. Additionally, HK2 appears to be important for preserving photoreceptors during aging. We show that retinal glucose metabolism is largely unchanged after HK2 deletion, suggesting that the non-enzymatic role of HK2 is important for maintaining photoreceptor health. These results suggest that HK2 expression is critical for preserving photoreceptors during acute nutrient stress and aging. More specifically, p-AKT mediated translocation of HK2 to the mitochondrial surface may be critical for protecting photoreceptors from acute and chronic stress.

Selective knockdown of hexokinase 2 in rods leads to age-related photoreceptor degeneration and retinal metabolic remodeling

“…We crossed RHO-Cre mice with HK2-floxed mice to selectively knock down HK2 in rods, with age-matched HK2 wild-type (WT) mice serving as controls. As previous studies found limited retinal changes around 5 months after disruption of glycolysis 13 , 14 , 19 , we characterized changes in the retina in two groups of mice: young mice at 19~20 weeks and aged mice 40~41 weeks of age. We used 4 retinas from individual mice per group for Western blots on each polyvinylidene difluoride (PVDF) membrane, 4 eyes/group for immunohistochemistry (IHC) on frozen sections, 8 mice/group for electroretinography (ERG), optical coherence tomography (OCT) and 7–8 retinas/group for metabolic analysis, in the study as described below.…”

“…HK1 is found in all mammalian tissues and considered as a “housekeeping enzyme” in physiological conditions. HK2 is the isoform that is highly expressed in photoreceptors 15 , 19 . HK1 and HK2 can associate with voltage-dependent-anion channel (VDAC) to regulate mitochondrial function 20 , 21 .…”

“…Little is known about the expression and function of HK3 and HK4 in the retina. Among the four isoforms of HK enzymes, HK2 plays a key role in the Warburg effect exhibited by photoreceptors 15 , 19 . Photoreceptors also express other key enzymes of glycolysis and the tricarboxylic acid (TCA) cycle, including pyruvate kinase and pyruvate dehydrogenase 13 , 15 , 22 , 23 .…”

“…It has been widely believed that photoreceptors utilize most glucose to generate lactate and only a small amount of pyruvate is metabolized through mitochondrial OXPHOS 3 , 15 , 18 , 25 . Recent studies have found that photoreceptors rely on glycolysis for the biogenesis of their outer segments and that genetic disruption of glycolysis results in photoreceptor degeneration 2 , 13 , 19 , 23 . However, Petit et al 14 recently proposed that aerobic glycolysis is not critical in photoreceptors but is rather a metabolic choice to maximize their function.…”

Phagocytosis by the Retinal Pigment Epithelium: Recognition, Resolution, Recycling

“…To do so, the RPE expresses remarkably high levels of GLUT1 at their apical and basolateral surface (164). Glucose in the cytosol would normally be quickly converted for its use in glycolysis, but this is effectively suppressed in the RPE by the lactate that is produced and exported by the photoreceptor and because kinases that act on sugars are inhibited or expressed at sufficient low levels (164,165). Conversely, such kinases (e.g.…”