Monitoring regional astrocyte diversity by cell type‐specific proteomic labeling in vivo

Abstract

Astrocytes exhibit regional heterogeneity in morphology, function and molecular composition to support and modulate neuronal function and signaling in a regionspecific manner. To characterize regional heterogeneity of astrocytic proteomes of different brain regions we established an inducible Aldh1l1-methionyl-tRNAsynthetase L274G (MetRS L274G ) mouse line that allows astrocyte-specific metabolic labeling of newly synthesized proteins by azidonorleucine (ANL) in vivo and subsequent isolation of tagged proteins by click chemistry. We analyzed astrocytic proteins from four different brain regions by mass spectrometry. The induced expression of MetRS L274G is restricted to astrocytes and identified proteins show a high overlap with proteins compiled in "AstroProt," a newly established database for astrocytic proteins. Gene enrichment analysis reveals a high similarity among brain regions with subtle differences in enriched biological processes and in abundances of key astrocytic proteins for hippocampus, cortex and striatum. However, the cerebellar proteome stands out with proteins being highly associated with the calcium signaling pathway or with bipolar disorder. Subregional analysis of single astrocyte TAMRA intensities in hippocampal layers indicates distinct subregional heterogeneity of astrocytes and highlights the applicability of our toolbox to study differences of astrocytic proteomes in vivo.

Cell-specific cross-talk proteomics reveals cathepsin B signaling as a driver of glioblastoma malignancy near the subventricular zone

“…Here, we use a nascent proteomic labeling system, MetRS L274G in the presence of ANL, to explore the proteomic contributions in the cross-talk between GBM and NPCs. Although this system has been used in other areas of neuroscience (62)(63)(64), cancer biology (65), and stem cell biology (66)(67)(68), neurogenesis and brain tumor cell-specific nascent proteomics has been relatively unexplored. Our approach to this system can now be used to profile GBM proteomic dynamics in vivo and can be more widely applied to other diseases and microenvironmental niches.…”

Altered calcium responses and antioxidant properties in Friedreich’s ataxia-like cerebellar astrocytes

“…This finding suggested that cerebellar astrocytes are more vulnerable to frataxin depletion than astrocytes from other brain regions. This specific cerebellar vulnerability could be partially attributed to the unique proteomic profile of astrocytes from the cerebellum, which differ from that of astrocytes in all other brain regions analyzed (Prabhakar et al, 2023). The cerebellum is characterized by astrocyte heterogeneity, resulting from a complex and still partially unclear process of glial cell proliferation and differentiation.…”

Advances in proteomic phenotyping of microglia in neurodegeneration

“…Because the BONCAT method only labels newly synthesized proteins, there may be challenges with achieving a desired depth of proteome, however, this feature may be an asset for studying event‐related or stimuli‐related changes to the proteome. Recently, BONCAT has been successfully applied to astrocytes as well [163]. The extension of the BONCAT method to label and profile microglia, other brain immune cells and glial subtypes has not been reported thus far.…”