Isoform Diversity and Regulation in Peripheral and Central Neurons Revealed through RNA-Seq

Abstract

To fully understand cell type identity and function in the nervous system there is a need to understand neuronal gene expression at the level of isoform diversity. Here we applied Next Generation Sequencing of the transcriptome (RNA-Seq) to purified sensory neurons and cerebellar granular neurons (CGNs) grown on an axonal growth permissive substrate. The goal of the analysis was to uncover neuronal type specific isoforms as a prelude to understanding patterns of gene expression underlying their intrinsic growth abilities. Global gene expression patterns were comparable to those found for other cell types, in that a vast majority of genes were expressed at low abundance. Nearly 18% of gene loci produced more than one transcript. More than 8000 isoforms were differentially expressed, either to different degrees in different neuronal types or uniquely expressed in one or the other. Sensory neurons expressed a larger number of genes and gene isoforms than did CGNs. To begin to understand the mechanisms responsible for the differential gene/isoform expression we identified transcription factor binding sites present specifically in the upstream genomic sequences of differentially expressed isoforms, and analyzed the 3′ untranslated regions (3′ UTRs) for microRNA (miRNA) target sites. Our analysis defines isoform diversity for two neuronal types with diverse axon growth capabilities and begins to elucidate the complex transcriptional landscape in two neuronal populations.

Palmitoylation controls DLK localization, interactions and activity to ensure effective axonal injury signaling

“…Taken together, these data suggest that DLK is palmitoylated in heterologous cells and in neurons. In contrast, homologous MAP3Ks that are expressed in sensory neurons (12) were not palmitoylated (Fig. S2).…”

“…Many of these DLK homologs are also expressed in peripheral neurons (12) and can activate JNK and p38 signaling in vitro and in transfected cells (13). These findings suggest that differential subcellular localization and regulation underlies DLK's unique role in nerve injury signaling, but the nature of any such DLK-specific regulation is unclear.…”

Beyond gene expression level: How are Bayesian methods doing a great job in quantification of isoform diversity and allelic imbalance?

“…Compared to microarrays, RNA expression profile has two majorly advantageous features to enable to identify complexity of isoform variability on a single gene to produce various splicing events on transcripts and another nature, allelic specific expression and allelic imbalance resulted from genetic differences in transcriptional rates (Beretta et al, 2014;Bernard et al, 2014;Deng et al, 2011;Hiller et al, 2009;Hiller and Wong, 2013;Howard and Heber, 2010;Hu et al, 2014;Jiang and Wong, 2009;Katz et al, 2010;Kaur et al, 2012;Kimes et al, 2014;Leon-Novelo et al, 2014;Lerch et al, 2012;Li and Jiang, 2012;Ma and Zhang, 2013;Mezlini et al, 2013;Mills et al, 2013;Nariai et al, 2013;Nariai et al, 2014;Ng et al, 2014;Nicolae et al, 2011;Niu et al, 2014;Pandey et al, 2013;Patro et al, 2014;Rehrauer et al, 2013;Safikhani et al, 2013;Shi and Jiang, 2013;Suo et al, 2014;Trapnell et al, 2010;Vardhanabhuti et al, 2013;Wang et al, 2010;Wu et al, 2011b;Yalamanchili et al, 2014;Zhang et al, 2014;Zheng and Chen, 2009). …”

How are Bayesian and Non-Parametric Methods Doing a Great Job in RNA-Seq Differential Expression Analysis? : A Review

“…In addition, the sample size in RNA-seq is much fewer than the number of variables (genes) (Anders et al, 2012;Beretta et al, 2014;Bernard et al, 2014;Bi and Davuluri, 2013;Bullard et al, 2010;Deng et al, 2011;Glaus et al, 2012;Han and Jiang, 2014;Hiller et al, 2009;Hiller and Wong, 2013;Howard and Heber, 2010;Hu et al, 2014;Jiang and Wong, 2009;Kaur et al, 2012;Kim et al, 2012;Kimes et al, 2014;Kumar et al, 2012;Lee et al, 2011;Leon-Novelo et al, 2014;Lerch et al, 2012;Li et al, 2014;Li et al, 2011;Li and Jiang, 2012;Ma and Zhang, 2013;Marioni et al, 2008;Mezlini et al, 2013;Mills et al, 2013;Mortazavi et al, 2008;Nariai et al, 2013;Nariai et al, 2014;Nicolae et al, 2011;Niu et al, 2014;Oh et al, 2013;Oshlack et al, 2010;Pandey et al, 2013;Patro et al, 2014;Pollier et al, 2013;Rehrauer et al, 2013;Roberts et al, 2011;Robinson and Oshlack, 2010;Ryan et al, 2012;Safikhani et al, 2013;<...>…”

“…And for an extreme case of gene, a gene has ∼ 1,000 isoform splicing events, suggesting that those isoforms with variability might be more closely related with disease and developmental processes as they generate different protein structure and functionalities, respectively (Beretta et al, 2014;Bernard et al, 2014;Deng et al, 2011;Hiller et al, 2009;Hiller and Wong, 2013;Howard and Heber, 2010;Hu et al, 2014;Jiang and Wong, 2009;Katz et al, 2010;Kaur et al, 2012;Kimes et al, 2014;Lerch et al, 2012;Li et al, 2011;Li and Jiang, 2012;Ma and Zhang, 2013;Mezlini et al, 2013;Mills et al, 2013;Nariai et al, 2013;Nariai et al, 2014;Nicolae et al, 2011;Niu et al, 2014;Patro et al, 2014;Rehrauer et al, 2013;Safikhani et al, 2013;Shi and Jiang, 2013;Suo et al, 2014;Trapnell et al, 2010;Vardhanabhuti et al, 2013;Wang et al, 2010b;Wang et al, 2010c;Wu et al, 2011;Yalamanchili et al, 2014;Zhang et al, 2014).…”