Transcriptome Profiling of Mouse Brain and Lung under Dip2A Regulation Using RNA-Sequencing

Abstract

Disconnected interacting 2 homolog A (DIP2A) gene is highly expressed in nervous system and respiratory system of developing embryos. However, genes regulated by Dip2a in developing brain and lung have not been systemically studied. Transcriptome of brain and lung in embryonic 19.5 day (E19.5) were compared between wild type and Dip2a-/- mice. Total RNAs were extracted from brain and lung of E19.5 embryos for RNA-Seq. Clean reads were mapped to mouse reference sequence (mm9) using Tophat and assembled into transcripts by Cufflinks. Edge R and DESeq were applied to identify differentially expressed genes (DEGs) and annotated under GO, COG, KEGG and TF. An average of 50 million reads per sample was mapped to the reference sequence. A total of 214 DEGs were detected in brain (82 up and 132 down) and 1900 DEGs in lung (1259 up and 641 down). GO enrichment analysis indicated that DEGs in both Brain and Lung were mainly enriched in biological processes ‘DNA-templated transcription and Transcription from RNA polymerase II promoter’, ‘multicellular organism development’, ‘cell differentiation’ and ‘apoptotic process’. In addition, COG classification showed that both were mostly involved in ‘Replication, Recombination and Repair’, ‘Signal transduction and mechanism’, ‘Translation, Ribosomal structure and Biogenesis’ and ‘Transcription’. KEGG enrichment analysis showed that brain was mainly enriched in ‘Thryoid cancer’ pathway whereas lung in ‘Complement and Coagulation Cascades’ pathway. Transcription factor (TF) annotation analysis identified Zinc finger domain containing (ZF) proteins were mostly regulated in lung and brain. Interestingly, study identified genes Skor2, Gpr3711, Runx1, Erbb3, Frmd7, Fut10, Sox11, Hapln1, Tfap2c and Plxnb3 from brain that play important roles in neuronal cell maturation, differentiation and survival; genes Hoxa5, Eya1, Ctsh, Erff1, Lama1, Lama2, Rspo2, Sox11, Spry4, Shh, Igf1 and Wnt7a from lung are important in lung development and morphogenesis. Expression levels of the candidate genes were validated by qRT-PCR. Genome wide transcriptional analysis using wild type and Dip2a knockout mice in brain and lung at embryonic day 19.5 (E19.5) provided a genetic basis of molecular function of these genes.