Epithelial (CD31-CD45-EpCAM+) cells were isolated by FACS from Grhl2-deficient (Shh-Cre;Grhl2f/-) and control (Shh-Cre;Grhl2f/+) embryonic lungs at day E16.5 (3 biological replicates/genotype). Total RNA extracted from the samples was subjected to next-generation sequencing (NGS) library preparation using standard Illumina protocols. Completed libraries from individual samples were sequenced on a HiSeq2500 at the Australian Genome Research Facility. Overall design: RNA-seq was performed on Grhl2-deficient and control epithelium isolated from the lungs of E16.5 embryos (n=3 replicates/genotype/cell population).
Lung morphogenesis is orchestrated through Grainyhead-like 2 (Grhl2) transcriptional programs.
Sex, Specimen part, Subject
View SamplesThe Krppel-like factors, KLF1 and KLF2, positively regulate embryonic -globin expression, and have additional overlapping roles in embryonic (primitive) erythropoiesis. KLF1-/-KLF2-/- double knockout mice are anemic at embryonic day 10.5 (E10.5) and die by E11.5, in contrast to single knockouts. To investigate the combined roles of KLF1 and KLF2 in primitive erythropoiesis, expression profiling of E9.5 erythroid cells was performed. A limited number of genes had a significantly decreasing trend of expression in wild-type, KLF1-/- and KLF1-/-KLF2-/-. Among these, c-myc emerged as a central node in the most significant gene network. c-myc expression is synergistically regulated by KLF1 and KLF2, and both factors bind the c-myc promoters. To characterize the role of c-myc in primitive erythropoiesis, ablation was performed specifically in mouse embryonic proerythroblast cells. After E9.5, these embryos exhibit an arrest in the normal expansion of circulating red cells and develop anemia analogous to KLF1-/-KLF2-/-. In the absence of c-myc, circulating erythroid cells do not show the normal increase in - and -like globin expression, but interestingly, have accelerated erythroid maturation, between E9.5 and E11.5. This study reveals a novel regulatory network by which KLF1 and KLF2 regulate c-myc, to control the primitive erythropoietic program.
Kruppel-like factor 1 (KLF1), KLF2, and Myc control a regulatory network essential for embryonic erythropoiesis.
Specimen part
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