RNA sequencing and proteomics approaches reveal novel deficits in the cortex of Mecp2-deficient mice, a model for Rett syndrome

Abstract

BackgroundRett syndrome (RTT) is an X-linked neurodevelopmental disorder caused by mutations in the transcriptional regulator MeCP2. Much of our understanding of MeCP2 function is derived from transcriptomic studies with the general assumption that alterations in the transcriptome correlate with proteomic changes. Advances in mass spectrometry-based proteomics have facilitated recent interest in the examination of global protein expression to better understand the biology between transcriptional and translational regulation.MethodsWe therefore performed the first comprehensive transcriptome-proteome comparison in a RTT mouse model to elucidate RTT pathophysiology, identify potential therapeutic targets, and further our understanding of MeCP2 function. The whole cortex of wild-type and symptomatic RTT male littermates (n = 4 per genotype) were analyzed using RNA-sequencing and data-independent acquisition liquid chromatography tandem mass spectrometry. Ingenuity® Pathway Analysis was used to identify significantly affected pathways in the transcriptomic and proteomic data sets.ResultsOur results indicate these two “omics” data sets supplement one another. In addition to confirming previous works regarding mRNA expression in Mecp2-deficient animals, the current study identified hundreds of novel protein targets. Several selected protein targets were validated by Western blot analysis. These data indicate RNA metabolism, proteostasis, monoamine metabolism, and cholesterol synthesis are disrupted in the RTT proteome. Hits common to both data sets indicate disrupted cellular metabolism, calcium signaling, protein stability, DNA binding, and cytoskeletal cell structure. Finally, in addition to confirming disrupted pathways and identifying novel hits in neuronal structure and synaptic transmission, our data indicate aberrant myelination, inflammation, and vascular disruption. Intriguingly, there is no evidence of reactive gliosis, but instead, gene, protein, and pathway analysis suggest astrocytic maturation and morphological deficits.ConclusionsThis comparative omics analysis supports previous works indicating widespread CNS dysfunction and may serve as a valuable resource for those interested in cellular dysfunction in RTT.Electronic supplementary materialThe online version of this article (10.1186/s13229-017-0174-4) contains supplementary material, which is available to authorized users.

Genome‐wide transcriptomic and proteomic studies of Rett syndrome mouse models identify common signaling pathways and cellular functions as potential therapeutic targets

“…In an attempt to identify region specific gene expression, studies were conducted on individual brain regions and cells isolated from these regions. These included the cortex (Kishi et al, ; Pacheco et al, ; Tudor, Akbarian, Chen, & Jaenisch, ; Urdinguio et al, ), visual cortex (Gabel et al, ; Renthal et al, ), the cerebellum (Ben‐Shachar et al, ; Gabel et al, ; Jordan, Li, Kwan, & Francke, ; Mellen, Ayata, & Heintz, ; Mellen, Ayata, Dewell, Kriaucionis, & Heintz, ; Sanfeliu, Hokamp, Gill, & Tropea, ; Urdinguio et al, ), whole brain (Guo et al, ; Kriaucionis et al, ; Nuber et al, ), the whole hippocampus (Baker et al, ; Tudor et al, ), and the amygdala (Samaco et al, ). Studies were also conducted on cell sources originating from distinct regions of the brain including forebrain (Tudor et al, ), hypothalamus (Chahrour et al, ; Chen et al, ), midbrain (Urdinguio et al, ), and striatum (Zhao, Goffin, Johnson, & Zhou, ), granule cells of the dentate gyrus hippocampal region (Smrt et al, ), embryonic cortical neurons (Bedogni et al, ; Vacca et al, ), and excitatory and inhibitory cortical neurons (Johnson et al, ).…”

“…The proteomic studies identified were conducted on three different Mecp2 mouse models: Mecp2 ‐null, Mecp2 308 , and Mecp2 Jae mice (Cortelazzo et al, ; Matarazzo & Ronnett, ; Pacheco et al, ). The ages of mice ranged from 2 weeks to 12‐months old.…”

“…The ages of mice ranged from 2 weeks to 12‐months old. Two studies were conducted on male mice (Matarazzo & Ronnett, ; Pacheco et al, ) with one study on female mice (Cortelazzo et al, ). Two studies were conducted on brain regions (Matarazzo & Ronnett, ; Pacheco et al, ) and one on plasma (Cortelazzo et al, ).…”

“…The three proteomic studies used slightly different protein analysis and quantification methods, with Matarazzo and Ronnett () using 2D gel electrophoresis and Q‐T of mass spectrometry. Cortelazzo et al () used 2D and MALDI‐ToF/ToF, while Pacheco et al () used an in‐gel tryptic digest and LC‐MS/MS.…”

Complex genetic network underlying the convergent of Rett Syndrome like (RTT-L) phenotype in neurodevelopmental disorders

“…As previously mentioned, RTT-L-causing genes are directly involved in the MECP2-involved pathways of chromatin regulation (HDAC2, MEF2C, CREB1, GRIN1) and upregulation of glutamate and downregulation of dopamine and GABA pathways (CREB1, KCNA2, GRIN1, KCNB1, GRIN2A, GRIN2B, and KCNQ2) 42 . And while the RTT-L-causing genes do not play a role in the transcriptome profiling described by Lin et al, they share overlapping biological processes in many neuronal functions that have altered expression in RTT, such as synaptic transmission, neurotransmission, excitation of neurons, and development of neurons 43,44 . The involvement of many RTT-L-implicated genes in MECP2 function indicates that the replication of many of clinical features of RTT-L similar to that of classical or atypical RTT could be attributed to the disruption of one of MECP2's transcription regulatory activities.…”