title: Targeted shRNA screening identified critical roles of pleckstrin-2 in erythropoiesis.
abstract: Differentiation of erythroblasts to mature red blood cells involves dynamic changes of the membrane and cytoskeleton networks that are not fully characterized. Using a mouse fetal liver erythroblast culture system and a targeted shRNA functional screening strategy, we identified a critical role of pleckstrin-2 in actin dynamics and protection of early stage terminal erythroblasts from oxidative damage. Knockdown of pleckstrin-2 in the early stage of terminal erythropoiesis disrupted the actin cytoskeleton and led to differentiation inhibition and apoptosis. This pro-survival and differentiation function of pleckstrin-2 was mediated through its interaction with cofilin, by preventing cofilin's mitochondrial entry when the intracellular level of reactive oxygen species was higher in the early stage of terminal erythropoiesis. Treatment of the cells with a scavenger of reactive oxygen species rescued cofilin's mitochondrial entry and differentiation inhibition induced by pleckstrin-2 knockdown. In contrast, pleckstrin-2 knockdown in late stage terminal erythroblasts had no effect on survival or differentiation but blocked enucleation due to disorganized actin cytoskeleton. Thus, our study identified a dual function of pleckstrin-2 in the early and late stages of terminal erythropoiesis through its regulations of actin dynamics and cofilin's mitochondrial localization, which reflects intracellular level of reactive oxygen species in different developmental stages.
authors.affiliation: Department of Pathology, Feinberg School of Medicine, Northwestern University, Chicago, IL.
authors.forename: Baobing
authors.initials: B
authors.lastname: Zhao
authors.affiliation: Department of Pathology, Feinberg School of Medicine, Northwestern University, Chicago, IL.
authors.forename: Ganesan
authors.initials: G
authors.lastname: Keerthivasan
authors.affiliation: Department of Pathology, Feinberg School of Medicine, Northwestern University, Chicago, IL.
authors.forename: Yang
authors.initials: Y
authors.lastname: Mei
authors.affiliation: Department of Pathology, Feinberg School of Medicine, Northwestern University, Chicago, IL.
authors.forename: Jing
authors.initials: J
authors.lastname: Yang
authors.affiliation: Department of Pathology, Feinberg School of Medicine, Northwestern University, Chicago, IL.
authors.forename: James
authors.initials: J
authors.lastname: McElherne
authors.affiliation: Whitehead Institute for Biomedical Research, Cambridge, MA.
authors.forename: Piu
authors.initials: P
authors.lastname: Wong
authors.affiliation: Broad Institute of Harvard University and the Massachusetts Institute of Technology, Cambridge, MA.
authors.forename: John G
authors.initials: JG
authors.lastname: Doench
authors.affiliation: Biomedical Informatics Center, Northwestern University, Chicago, IL, USA.
authors.forename: Gang
authors.initials: G
authors.lastname: Feng
authors.affiliation: Broad Institute of Harvard University and the Massachusetts Institute of Technology, Cambridge, MA.
authors.forename: David E
authors.initials: DE
authors.lastname: Root
authors.affiliation: Department of Pathology, Feinberg School of Medicine, Northwestern University, Chicago, IL peng-ji@fsm.northwestern.edu.
authors.forename: Peng
authors.initials: P
authors.lastname: Ji
mesh_terms: D051339:Actin Depolymerizing Factors / Q000378:metabolism; D000199:Actins / Q000235:genetics / Q000378:metabolism; D000818:Animals; D017209:Apoptosis / Q000235:genetics; D002454:Cell Differentiation / Q000235:genetics; D004900:Erythroblasts / Q000166:cytology / Q000378:metabolism; D004920:Erythropoiesis / Q000235:genetics*; D020869:Gene Expression Profiling; D005786:Gene Expression Regulation; D017930:Genes, Reporter; D008565:Membrane Proteins / Q000235:genetics* / Q000378:metabolism; D051379:Mice; D008928:Mitochondria / Q000235:genetics / Q000378:metabolism; D011485:Protein Binding; D034622:RNA Interference; D034741:RNA, Small Interfering / Q000235:genetics*
chemical_list: D051339:Actin Depolymerizing Factors; D000199:Actins; D008565:Membrane Proteins; C485635:Plek2 protein, mouse; D034741:RNA, Small Interfering
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