title: Knockdown of prothymosin α leads to apoptosis and developmental defects in zebrafish embryos.
abstract: Prothymosin alpha (ProTα) is an abundant nuclear protein involved in cellular processes intricately linked to development, such as cell proliferation and apoptosis. Although it is known that ProTα inhibits the formation of apoptosome and blocks caspase-3 activity, its mechanism of function in the apoptotic machinery is still under investigation. We have studied the cellular role of ProTα by knocking down its expression in HeLa cells with small hairpin RNA (shRNA) in the absence of apoptotic stimuli. Flow cytometric analysis showed that the live cell population was significantly decreased with a concomitant increase of the apoptotic populations. To understand the physiological role of ProTα within the context of embryonic development, we knocked down the Ptmab zebrafish ortholog using 2 specific morpholino oligonucleotides. Ptmab morphants exhibited growth retardation, bended trunks, and curly tails. The frequency of occurrence of the phenotypic defects was increased in a morpholino dose-dependent manner. Co-injection of ptmaa mRNA with ptmab morpholino partially rescued the morphological defects. Immunostaining with the anti-phospho-histone H3 (pH3) antibody suggested that the abnormalities of Ptmab morphants could be due to defective cell proliferation that results in growth imbalances. TUNEL fluorescent labelling and Acridine Orange staining of the morphants showed high rates of cell death in the head and tail regions. Concomitantly, the active form of caspase-3 was detected in Ptmab morphants. Our data suggest a conserved anti-apoptotic role of ProTα between zebrafish and humans, and provide the first evidence that ProTα is important for early embryogenesis.
authors.affiliation: a Laboratory of Biological Chemistry, Medical School, University of Ioannina, 451 10 Ioannina, Greece.
authors.forename: Anastasia
authors.initials: A
authors.lastname: Emmanouilidou
authors.forename: Zoe
authors.initials: Z
authors.lastname: Karetsou
authors.forename: Eleni
authors.initials: E
authors.lastname: Tzima
authors.forename: Takahiko
authors.initials: T
authors.lastname: Kobayashi
authors.forename: Thomais
authors.initials: T
authors.lastname: Papamarcaki
mesh_terms: D000818:Animals; D017209:Apoptosis / Q000235:genetics*; D053148:Caspase 3 / Q000378:metabolism; D002460:Cell Line; D049109:Cell Proliferation; D004625:Embryo, Nonmammalian / Q000166:cytology / Q000196:embryology / Q000378:metabolism; D018507:Gene Expression Regulation, Developmental; D006367:HeLa Cells; D006801:Humans; D060172:Morpholinos / Q000235:genetics; D011498:Protein Precursors / Q000235:genetics / Q000378:metabolism*; D034622:RNA Interference; D012333:RNA, Messenger / Q000235:genetics; D034741:RNA, Small Interfering; D013947:Thymosin / Q000031:analogs & derivatives* / Q000235:genetics / Q000378:metabolism; D015027:Zebrafish / Q000002:abnormalities* / Q000235:genetics
chemical_list: D060172:Morpholinos; D011498:Protein Precursors; D012333:RNA, Messenger; D034741:RNA, Small Interfering; C040782:prothymosin alpha; D013947:Thymosin; D053148:Caspase 3