mTORC1 regulates high levels of protein synthesis in retinal ganglion cells of adult mice.

Mechanistic target of rapamycin (mTOR) and mTOR complex 1 (mTORC1), linchpins of the nutrient sensing and protein synthesis pathways, are present at relatively high levels in the ganglion cell layer (GCL) and retinal ganglion cells (RGCs) of rodent and human retinas. However, the role of mTORCs in the control of protein synthesis in RGC is unknown. Here, we applied the SUrface SEnsing of Translation (SUnSET) method of nascent protein labeling to localize and quantify protein synthesis in the retinas of adult mice.

Insulin-like growth factor-2 regulates basal retinal insulin receptor activity.

The retinal insulin receptor (IR) exhibits basal kinase activity equivalent to that of the liver of fed animals, but unlike the liver, does not fluctuate with feeding and fasting; it also declines rapidly after the onset of insulin-deficient diabetes. The ligand(s) that determine basal IR activity in the retina has not been identified. Using a highly sensitive insulin assay, we found that retinal insulin concentrations remain constant in fed versus fasted rats and in diabetic versus control rats; vitreous fluid insulin levels were undetectable.

mTORC1 and mTORC2 expression in inner retinal neurons and glial cells.

The retina is one of the most metabolically active tissues, yet the processes that control retinal metabolism remains poorly understood. The mTOR complex (mTORC) that drives protein and lipid biogenesis and autophagy has been studied extensively in regards to retinal development and responses to optic nerve injury but the processes that regulate homeostasis in the adult retina have not been determined. We previously demonstrated that normal adult retina has high rates of protein synthesis compared to skeletal muscle, associated with high levels of mechanistic target of rapamycin (mTOR), a kinase that forms multi-subunit complexes that sense and integrate diverse environmental cues to control cell and tissue physiology.

Phosphatase control of 4E-BP1 phosphorylation state is central for glycolytic regulation of retinal protein synthesis.

Control of protein synthesis in insulin-responsive tissues has been well characterized, but relatively little is known about how this process is regulated in nervous tissues. The retina exhibits a relatively high protein synthesis rate, coinciding with high basal Akt and metabolic activities, with the majority of retinal ATP being derived from aerobic glycolysis. We examined the dependency of retinal protein synthesis on the Akt-mTOR signaling and glycolysis using ex vivo rat retinas.

mTORC1-independent reduction of retinal protein synthesis in type 1 diabetes.

Poorly controlled diabetes has long been known as a catabolic disorder with profound loss of muscle and fat body mass resulting from a simultaneous reduction in protein synthesis and enhanced protein degradation. By contrast, retinal structure is largely maintained during diabetes despite reduced Akt activity and increased rate of cell death. Therefore, we hypothesized that retinal protein turnover is regulated differently than in other insulin-sensitive tissues, such as skeletal muscle.

Differential roles of hyperglycemia and hypoinsulinemia in diabetes induced retinal cell death: evidence for retinal insulin resistance.

Diabetes pathology derives from the combination of hyperglycemia and hypoinsulinemia or insulin resistance leading to diabetic complications including diabetic neuropathy, nephropathy and retinopathy. Diabetic retinopathy is characterized by numerous retinal defects affecting the vasculature and the neuro-retina, but the relative contributions of the loss of retinal insulin signaling and hyperglycemia have never been directly compared. In this study we tested the hypothesis that increased retinal insulin signaling and glycemic normalization would exert differential effects on retinal cell survival and retinal physiology during diabetes.

Diabetes impairs the neuroprotective properties of retinal alpha-crystallins.

Purpose: α-Crystallins are small heat shock proteins that regulate cellular damage and cell survival. Expression of the proteins of the crystallin superfamily in the retina and their role in neuronal cell survival were investigated in two animal models of diabetes and retinal neurons in culture.

Methods: Crystallin expression was assessed in streptozotocin-induced and Ins2(Akita) diabetic mice using iTRAQ methodology and validated using immunoblotting.

Genetic evidence for sites of interaction between the Gal3 and Gal80 proteins of the Saccharomyces cerevisiae GAL gene switch.

Galactose-activated transcription of the Saccharomyces cerevisiae GAL genes occurs when Gal3 binds the Gal4 inhibitor, Gal80. Noninteracting variants of Gal3 or Gal80 render the GAL genes noninducible. To identify the binding determinants for Gal3's interaction with Gal80 we carried out GAL3-GAL80 intergenic suppression analyses and selected for new GAL3 mutations that impair the Gal3-Gal80 interaction.

Regulation of T cell development by the deubiquitinating enzyme CYLD.

T cell receptor signaling is essential for the generation and maturation of T lymphocyte precursors. Here we identify the deubiquitinating enzyme CYLD as a positive regulator of proximal T cell receptor signaling in thymocytes. CYLD physically interacted with active Lck and promoted recruitment of active Lck to its substrate, Zap70.