ABSTRACT
Hedgehog (Hh) signaling relies on the primary cilium, a cell surface organelle that serves as a signaling hub for the cell. Using proximity labeling and quantitative proteomics, we identify Numb as a ciliary protein that positively regulates Hh signaling. Numb localizes to the ciliary pocket and acts as an endocytic adaptor to incorporate Ptch1 into clathrin-coated vesicles, thereby promoting Ptch1 exit from the cilium, a key step in Hh signaling activation. Numb loss impedes Sonic hedgehog (Shh)-induced Ptch1 exit from the cilium, resulting in reduced Hh signaling. Numb loss in spinal neural progenitors reduces Shh-induced differentiation into cell fates reliant on high Hh activity. Genetic ablation of Numb in the developing cerebellum impairs the proliferation of granule cell precursors, a Hh-dependent process, resulting in reduced cerebellar size. This study highlights Numb as a regulator of ciliary Ptch1 levels during Hh signal activation and demonstrates the key role of ciliary pocket-mediated endocytosis in cell signaling.
Subject(s)
Cerebellum , Cilia , Hedgehog Proteins , Nerve Tissue Proteins , Patched-1 Receptor , Signal Transduction , Hedgehog Proteins/metabolism , Hedgehog Proteins/genetics , Cilia/metabolism , Animals , Patched-1 Receptor/metabolism , Patched-1 Receptor/genetics , Mice , Nerve Tissue Proteins/metabolism , Nerve Tissue Proteins/genetics , Cerebellum/metabolism , Membrane Proteins/metabolism , Membrane Proteins/genetics , Humans , Endocytosis , Cell Differentiation , Cell Proliferation , Neural Stem Cells/metabolism , Neural Stem Cells/cytology , Mice, KnockoutABSTRACT
Neural progenitor cells within the cerebral cortex undergo a characteristic switch between symmetric self-renewing cell divisions early in development and asymmetric neurogenic divisions later. Yet, the mechanisms controlling this transition remain unclear. Previous work has shown that early but not late neural progenitor cells (NPCs) endogenously express the autism-linked transcription factor Foxp1, and both loss and gain of Foxp1 function can alter NPC activity and fate choices. Here, we show that premature loss of Foxp1 upregulates transcriptional programs regulating angiogenesis, glycolysis, and cellular responses to hypoxia. These changes coincide with a premature destabilization of HIF-1α, an elevation in HIF-1α target genes, including Vegfa in NPCs, and precocious vascular network development. In vitro experiments demonstrate that stabilization of HIF-1α in Foxp1-deficient NPCs rescues the premature differentiation phenotype and restores NPC maintenance. Our data indicate that the endogenous decline in Foxp1 expression activates the HIF-1α transcriptional program leading to changes in the tissue environment adjacent to NPCs, which, in turn, might alter their self-renewal and neurogenic capacities.
Subject(s)
Cerebral Cortex , Forkhead Transcription Factors , Hypoxia-Inducible Factor 1, alpha Subunit , Neural Stem Cells , Repressor Proteins , Signal Transduction , Hypoxia-Inducible Factor 1, alpha Subunit/metabolism , Hypoxia-Inducible Factor 1, alpha Subunit/genetics , Forkhead Transcription Factors/metabolism , Forkhead Transcription Factors/genetics , Neural Stem Cells/metabolism , Neural Stem Cells/cytology , Animals , Mice , Cerebral Cortex/metabolism , Cerebral Cortex/cytology , Repressor Proteins/metabolism , Repressor Proteins/genetics , Neovascularization, Physiologic/genetics , Cell Differentiation/genetics , Vascular Endothelial Growth Factor A/metabolism , Vascular Endothelial Growth Factor A/genetics , Neurogenesis/genetics , Glycolysis , AngiogenesisABSTRACT
OBJECTIVE: Undernutrition has been associated with gut mucosal atrophy, impaired absorption, and increased permeability. This study investigated the effect of severe undernutrition and subsequent refeeding on gastric and duodenal mucosal protein fractional syntheses (MPFS). METHODS: MPFS was investigated in the gastric body, antrum, and duodenum of 23 severely undernourished patients by means of (14)C-leucine incorporation into tissue protein and repeated after a period of refeeding. Results were evaluated in comparison with a group of 22 healthy controls and presented as mean +/- standard error of the mean. RESULTS: The mean body index of the undernourished patients was 13.29 +/- 0.33 kg/m(2) versus 22.67 +/- 0.63 kg/m(2) in the controls (P < 0.001). MPFS in the controls and undernourished patients were similar (gastric body, 36.49 +/- 2.88 versus 33.41 +/- 3.08%/d; gastric antrum, 25.51 +/- 2.20 versus 24.95 +/- 2.32%/d; duodenum, 25.90 +/- 2.58 versus 25.49 +/- 1.99%/d). After refeeding, the body mass index of the undernourished patients increased to 15.87 +/- 0.44 kg/m(2) (P < 0.001). The MPFS increased significantly (gastric body, 51.80 +/- 8.12%/d, P < 0.05; gastric antrum, 33.44 +/- 3.66%/d, P < 0.05; duodenum, 46.27 +/- 8.02%/d, P < 0.01), with the MPFS of the duodenum significantly greater than the control values (P = 0.01). CONCLUSION: Despite severe undernutrition, MPFS of the gastric body, antrum, and duodenum remained similar to control values. Enteral feeding resulted in a significant increase in MPFS, indicating a trophic response.
