Neuronal Differentiation of LUHMES Cells Induces Substantial Changes of the Proteome.

Neuronal cell lines are important model systems to study mechanisms of neurodegenerative diseases. One example is the Lund Human Mesencephalic (LUHMES) cell line, which can differentiate into dopaminergic-like neurons and is frequently used to study mechanisms of Parkinson's disease and neurotoxicity. Neuronal differentiation of LUHMES cells is commonly verified with selected neuronal markers, but little is known about the proteome-wide protein abundance changes during differentiation. Using mass spectrometry and label-free quantification (LFQ), the proteome of differentiated and undifferentiated LUHMES cells and of primary murine midbrain neurons are compared. Neuronal differentiation induced substantial changes of the LUHMES cell proteome, with proliferation-related proteins being strongly down-regulated and neuronal and dopaminergic proteins, such as L1CAM and α-synuclein (SNCA) being up to 1,000-fold up-regulated. Several of these proteins, including MAPT and SYN1, may be useful as new markers for experimentally validating neuronal differentiation of LUHMES cells. Primary midbrain neurons are slightly more closely related to differentiated than to undifferentiated LUHMES cells, in particular with respect to the abundance of proteins related to neurodegeneration. In summary, the analysis demonstrates that differentiated LUHMES cells are a suitable model for studies on neurodegeneration and provides a resource of the proteome-wide changes during neuronal differentiation. (ProteomeXchange identifier PXD020044).

An optimized quantitative proteomics method establishes the cell type-resolved mouse brain secretome.

To understand how cells communicate in the nervous system, it is essential to define their secretome, which is challenging for primary cells because of large cell numbers being required. Here, we miniaturized secretome analysis by developing the "high-performance secretome protein enrichment with click sugars" (hiSPECS) method. To demonstrate its broad utility, hiSPECS was used to identify the secretory response of brain slices upon LPS-induced neuroinflammation and to establish the cell type-resolved mouse brain secretome resource using primary astrocytes, microglia, neurons, and oligodendrocytes. This resource allowed mapping the cellular origin of CSF proteins and revealed that an unexpectedly high number of secreted proteins in vitro and in vivo are proteolytically cleaved membrane protein ectodomains. Two examples are neuronally secreted ADAM22 and CD200, which we identified as substrates of the Alzheimer-linked protease BACE1. hiSPECS and the brain secretome resource can be widely exploited to systematically study protein secretion and brain function and to identify cell type-specific biomarkers for CNS diseases.