FAM69C functions as a kinase for eIF2α and promotes stress granule assembly.

Stress granules are dynamic cytoplasmic ribonucleoprotein granules that assemble in response to cellular stress. Aberrant formation of stress granules has been linked to neurodegenerative diseases. However, the molecular mechanisms underlying the initiation of stress granules remain elusive. Here we report that the brain-enriched protein kinase FAM69C promotes stress granule assembly through phosphorylation of eukaryotic translation initiation factor 2 (eIF2α). FAM69C physically interacts with eIF2α and functions as a stress-specific kinase for eIF2α, leading to stress-induced protein translation arrest and stress granule assembly. Primary microglia derived from Fam69c knockout mice exhibit aberrant stress granule assembly in response to oxidative stress and ATP. Defective stress granule assembly in microglia correlates with the formation of ASC specks and NLRP3 inflammasome activation, whereas induction of stress granule precludes inflammasome formation. Consistently, increased NLRP3 levels, caspase-1 cleavage and Il18 expression corroborate microglia-associated neuroinflammation in aged Fam69c knockout mice. Our study demonstrates that FAM69C is critical for stress granule assembly and suggests its role in the regulation of microglia function.

FAM69C, a kinase critical for synaptic function and memory, is defective in neurodegenerative dementia.

Synapse loss and memory decline are the primary features of neurodegenerative dementia. However, the molecular underpinnings that drive memory loss remain largely unknown. Here, we report that FAM69C is a kinase critically involved in neurodegenerative dementia. Biochemical analyses uncover that FAM69C is a serine/threonine kinase. We generate the Fam69c knockout mice and show by single-cell RNA sequencing that FAM69C deficiency drives cell-type-specific transcriptional changes relevant to synapse dysfunction. Electrophysiological, morphological, and behavioral experiments demonstrate impairments in synaptic plasticity, dendritic spine density, and memory in Fam69c knockout mice, as well as stress-induced neuronal death. Phosphoproteomic characterizations reveal that FAM69C substrates are involved in synaptic structure and function. Finally, reduced levels of FAM69C are found in postmortem brains of Alzheimer's disease patients. Our study demonstrates that FAM69C is a protective regulator of memory and suggests FAM69C as a potential therapeutic target for memory loss in neurodegenerative dementia.

Facile preparation of 3-D floor-like ordered mesoporous carbon functionalized graphene composites and its application for selective enrichment of N-glycans from human serum.

Novel 3-D floor-like ordered mesoporous carbon functionalized graphene composites, FLOMC-GO, with high graphitized carbon contents were successfully synthesized using a soft template method. The one-pot sol-gel method was employed to prepare the silica soft template. Then, the sandwich-like composites were further combined together to form a 3-D structure through pre-carbonization and carbonization. During these procedures, the sulphonyl and sulfide bridges were formed by cross-linking processes to connect the phenyl rings. The prepared FLOMC-GO was confirmed to have a large pore volume (1.03cm3g-1), high BET surface area (544.99m2g-1) and well-ordered mesoporous structure with a narrow pore-size concentrated at 3.74nm. The content of carbon reached 80% and was highly graphitized. By taking advantage of the interactions between carbon and glycans, FLOMC-GO was utilized to enrich N-linked glycans from OVA and human serum. As expected, excellent size-exclusion was found during the enrichment of N-glycans released from OVA, and 25 N-linked glycans were identified. The intensity of glycans enriched by FLOMC-GO was 7 times to the result of active carbon, while the ratio of OVA digestion to BSA interfering proteins increased to 1:50. Additionally, 31 N-linked glycans in total were enriched from human serum. The relatively easy synthesis as well as ability to enrich N-linked glycans with high selectivity and efficiency makes FLOMC-GO a promising adsorbent material for the discovery of human serum biomarkers for disease diagnosis.

Synthesis of magnetic zwitterionic-hydrophilic material for the selective enrichment of N-linked glycopeptides.

In consideration of the close connection between glycopeptides and human diseases, the efficient method to separate and enrich glycopeptides from complex biological samples is urgently required. In the work, we developed a magnetic zwitterionic-hydrophilic material for highly effective separation and analysis of glycopeptides from complex samples. The Fe3O4 particles were covered with a thick layer of polymer by one-step reflux-precipitation polymerization (RPP), subsequently decorated by Au nanoparticles (Au NPs) through in situ reduction and finally modified with zwitterionic groups. The abundant zwitterionic sites facilitate the selective enrichment of glycopeptides. Besides, the prepared Fe3O4@PGMA@Au-l-cys showed high detection sensitivity (5 fmol IgG digest), approving enrichment capacity (75mgg-1), satisfactory enrichment recovery (89.8%), and great performance in the analysis and profiling of low-abundance N-linked glycopeptides. Furthermore, the prepared material was employed in the enrichment of glycopeptides in intricate biological samples, and 774 unique N-glycosylation sites from 411 N-glycosylated proteins were reliably identified in three replicate analyses of a 75µg protein sample extracted from mouse liver, suggesting wide application prospect in glycoproteomics.

Selective extraction of endogenous peptides from human serum with magnetic mesoporous carbon composites.

Highly sensitive and selective enrichment of endogenous peptides or proteins from complex bio-system takes a significant important place to the proteomic. In this work, a unique Fe3O4@2SiO2@mSiO2-C nanomaterial was synthesized, contributing to the separation and enrichment of low concentration peptides from complex mixture. The highly ordered mesoporous carbon structure render the nanospheres with unique properties of strongly connected pore channels, strong hydrophobic properties, high specific surface area (254.90m2/g), uniform pore size (3.61nm). Which made it a promising candidates for the efficient enrichment of peptides through hydrophobic-hydrophobic interaction with low detection limit (0.2fmol), superb size-exclusion of high molecular weight proteins, highly selectivity for BSA digest (molar ratio of BSA tryptic digests/BSA, 1:400), ideal peptides recovery (about 87.5%), wonderful repeatability (RSD less than 25%). Moreover, the as-prepared Fe3O4@2SiO2@mSiO2-C nanoparticles were successfully enriched 2198 endogenous peptides from human serum, which fully indicated that the mesoporous carbon nanoparticles was a promising candidate for isolating proteins or peptides from complex biologicals.

Facile Preparation of Core-Shell Magnetic Metal-Organic Framework Nanoparticles for the Selective Capture of Phosphopeptides.

In regard to the phosphoproteome, highly specific and efficient capture of heteroideous kinds of phosphopeptides from intricate biological sample attaches great significance to comprehensive and in-depth phosphorylated proteomics research. However, until now, it has been a challenge. In this study, a new-fashioned porous immobilized metal ion affinity chromatography (IMAC) material was designed and fabricated to promote the selectivity and detection limit for phosphopeptides by covering a metal-organic frameworks (MOFs) shell onto Fe3O4 nanoparticles, taking advantage of layer-by-layer method (the synthesized nanoparticle denoted as Fe3O4@MIL-100 (Fe)). The thick layer renders the nanoparticles with perfect hydrophilic character, super large surface area, large immobilization of the Fe(3+) ions and the special porous structure. Specifically, the as-synthesized MOF-decorated magnetic nanoparticles own an ultra large surface area which is up to 168.66 m(2) g(-1) as well as two appropriate pore sizes of 1.93 and 3.91 nm with a narrow grain-size distribution and rapid separation under the magnetic circumstance. The unique features vested the synthesized nanoparticles an excellent ability for phosphopeptides enrichment with high selectivity for ß-casein (molar ratio of ß-casein/BSA, 1:500), large enrichment capacity (60 mg g(-1)), low detection limit (0.5 fmol), excellent phosphopeptides recovery (above 84.47%), fine size-exclusion of high molecular weight proteins, good reusability, and desirable batch-to-batch repeatability. Furthermore, encouraged by the experimental results, we successfully performed the as-prepared porous IMAC nanoparticle in the specific capture of phosphopeptides from the human serum (both the healthy and unhealthy) and nonfat milk, which proves itself to be a good candidate for the enrichment and detection of the low-abundant phosphopeptides from complicated biological samples.