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Proteomic analysis of FPA interactors

ABSTRACT: Genes involved in disease resistance are some of the fastest evolving and most diverse components of genomes. Large numbers of nucleotide-binding, leucine-rich repeat receptor genes (NLRs) are found in plant genomes and provide disease resistance. However, NLRs can trigger autoimmunity, disrupt beneficial microbiota or reduce fitness. It is therefore crucial to understand how NLRs are controlled. Here we show that the RNA-binding protein FPA mediates widespread premature cleavage and polyadenylation of NLR transcripts, controlling their functional expression and impacting immunity. Using long-read nanopore direct RNA sequencing we resolved the complexity of NLR transcript processing and gene annotation. Our results uncover a co-transcriptional layer of NLR control with implications for understanding the regulatory and evolutionary dynamics of NLRs in immunity.

INSTRUMENT(S): LTQ Orbitrap Velos

ORGANISM(S): Arabidopsis Thaliana (mouse-ear Cress)

SUBMITTER: Matthew Parker

LAB HEAD: Gordon Simpson

PROVIDER: PXD022684 | Pride | 2020-12-16

REPOSITORIES: Pride

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Project description:The winged helix protein FOXA2 and the nuclear receptor PPARg are highly conserved, regionally-expressed transcription factors that regulate networks of genes controlling complex metabolic functions. Cistrome analysis for FOXA2 in mouse liver and PPARg in mouse adipocytes has previously produced consensus binding sites that are nearly identical to those used by the factors in human cells. Despite this conservation of the canonical binding motif, we report here that the great majority of specific binding regions for FOXA2 in human liver and for PPARg in human adipocytes are not in the orthologous locations to the mouse genome. Nevertheless, gene-centric analysis reveals strong shared transcription factor occupancy near genes in tissue-specific metabolic pathways that are functionally conserved across species. Genes with only species-specific binding sites fail to show enrichment for these pathways. Thus, the biological functions of transcription factors that control specific metabolic functions are highly shared across species. Two TFs, FOXA2 and PPARg, were studied for genome-wide conservation of binding between mouse and human in specific tissues/cell-types (liver for FOXA2, adipocytes for PPARg). The number of replicates for each TF was chosen to obtain a comparable number of reads between the TFs and species. Human FOXA2 ChIP-seq was performed on two biological replicates of human liver samples, in three technical replicates each. Input DNA was also collected and sequenced from both biological samples. Mouse FOXA2 ChIP-seq was performed on four biological replicates of mouse liver samples. The ChIP and sequencing were repeated on two of these biological replicates to create technical replicates for additional sequence reads. Input DNA was sequenced from three additional mouse livers. Human PPARg ChIP-seq was performed on a human adipocyte cell-line (SGBS) differentiated in two replicate cultures. Input DNA was also collected and sequenced from one culture. Mouse PPARg ChIP-seq was performed on 3T3-L1 cells differentiated into adipocytes in culture in a single replicate, and this sequence data was pooled with existing data previously generated by the same lab, already available in GEO (GSE21314). A standard pool of input DNA sample sequence from multiple mouse tissue was used for analyzing the Mouse PPARg ChIP-seq data.

			Action	DRS
	Araport11_genes.201606.pep.repr.fasta	Fasta
	FPA_1_1.raw	Raw
	FPA_1_2.raw	Raw
	FPA_1_3.raw	Raw
	FPA_1_4.raw	Raw

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Proteomic analysis of FPA interactors

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