Project description:The properties of the cell types that are most vulnerable in the Huntington's disease (HD) cortex have not been delineated. Here we have employed serial fluorescence activated nuclear sorting (sFANS) and deep molecular profiling to demonstrate that layer 5a pyramidal neurons are selectively vulnerable in primary motor cortex and other cortical areas.
Project description:To study the properties of the major cell types of human cortex we have employed serial fluorescence activated nuclear sorting (sFANS) and deep molecular profiling.
Project description:To reveal gene expression programs altered in the deep layer neuron cell classes of Huntington's Disease samples, we first filtered the cell-type specific differential gene expression lists from our DESeq2 analyses based on the cell-type specific accessible promotor peaks revealed through ATACseq of control deep layer nuclei. This step allowed us to focus our gene lists for the following GO analyses to those genes that were truly accessible in the specific cell-types.
Project description:We sought to more precisely characterize the different alpha-synuclein (aSyn) 3’UTR mRNA species in normal and PD human brain. High-throughput, whole-transcriptome sequencing of the 3’UTR ends of polyadenylated mRNA transcripts (termed pA-RNAseq; see Methods) was performed on a cohort of 17 unaffected and 17 PD cerebral cortical tissue samples. This revealed 5 aSyn 3’UTR isoforms, with lengths of 290, 480, 560, 1070 and 2520 nt. Of these, the 560 nt and 2520 nt forms were predominant. The existence and relative preponderance of these species was further confirmed by Northern Blot. We next hypothesized, that aSyn 3’UTR selection might be altered in PD. Comparison of pA-RNAseq profiles from PD and unaffected cerebral cortex samples revealed an increase in the preponderance of the long 3’UTR species (>560 nt) relative to shorter species (<560 nt). Such a relative increase in aSynL was confirmed by Quantitative real-time RT-PCR (rt-qPCR) and appeared specific for PD, as the increase was also observed by comparison to RNA from amyotrophic lateral sclerosis patient samples. We note that the modified aSyn 3’UTR selection associated with PD patient tissue was detected in cerebral cortex tissue, which typically harbors pathological evidence of the disease process without frank cell loss; thus, this phenotype is unlikely to be a secondary consequence of neurodegeneration.
Project description:We sought to more precisely characterize the different alpha-synuclein (aSyn) 3M-bM-^@M-^YUTR mRNA species in normal and PD human brain. High-throughput, whole-transcriptome sequencing of the 3M-bM-^@M-^YUTR ends of polyadenylated mRNA transcripts (termed pA-RNAseq; see Methods) was performed on a cohort of 17 unaffected and 17 PD cerebral cortical tissue samples. This revealed 5 aSyn 3M-bM-^@M-^YUTR isoforms, with lengths of 290, 480, 560, 1070 and 2520 nt. Of these, the 560 nt and 2520 nt forms were predominant. The existence and relative preponderance of these species was further confirmed by Northern Blot. We next hypothesized, that aSyn 3M-bM-^@M-^YUTR selection might be altered in PD. Comparison of pA-RNAseq profiles from PD and unaffected cerebral cortex samples revealed an increase in the preponderance of the long 3M-bM-^@M-^YUTR species (>560 nt) relative to shorter species (<560 nt). Such a relative increase in aSynL was confirmed by Quantitative real-time RT-PCR (rt-qPCR) and appeared specific for PD, as the increase was also observed by comparison to RNA from amyotrophic lateral sclerosis patient samples. We note that the modified aSyn 3M-bM-^@M-^YUTR selection associated with PD patient tissue was detected in cerebral cortex tissue, which typically harbors pathological evidence of the disease process without frank cell loss; thus, this phenotype is unlikely to be a secondary consequence of neurodegeneration. Comparison of 3'UTR ends of alpha-synuclein in PD and unaffected brain cortex
Project description:The properties of the cell types that are most vulnerable in the Huntington’s disease (HD) cortex have not been delineated. Here we have employed serial fluorescence activated nuclear sorting (sFANS) and deep molecular profiling to demonstrate that layer 5a pyramidal neurons are selectively vulnerable in primary motor cortex and other cortical areas.