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TopBP1 Governs Hematopoietic Stem/Progenitor Cells Survival in Zebrafish Definitive Hematopoiesis.

Gao L, Li D, Ma K, Zhang W, Xu T, Fu C, Jing C, Jia X, Wu S, Sun X, Dong M, Deng M, Chen Y, Zhu W, Peng J, Wan F, Zhou Y, Zon LI, Pan W - PLoS Genet. (2015)

Bottom Line: Homozygous topbp1cas003 mutants manifest reduced number of HSPCs during definitive hematopoietic cell expansion, without affecting the formation and migration of HSPCs.Moreover, HSPCs in the caudal hematopoietic tissue (an equivalent of the fetal liver in mammals) in topbp1cas003 mutant embryos are more sensitive to hydroxyurea (HU) treatment.Collectively, this study demonstrates a novel and vital role of TopBP1 in the maintenance of HSPCs genome integrity and survival during hematopoietic progenitor expansion.

View Article: PubMed Central - PubMed

Affiliation: Key Laboratory of Stem Cell Biology, Institute of Health Sciences, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences & Shanghai Jiao Tong University School of Medicine, Shanghai, China.

ABSTRACT
In vertebrate definitive hematopoiesis, nascent hematopoietic stem/progenitor cells (HSPCs) migrate to and reside in proliferative hematopoietic microenvironment for transitory expansion. In this process, well-established DNA damage response pathways are vital to resolve the replication stress, which is deleterious for genome stability and cell survival. However, the detailed mechanism on the response and repair of the replication stress-induced DNA damage during hematopoietic progenitor expansion remains elusive. Here we report that a novel zebrafish mutantcas003 with nonsense mutation in topbp1 gene encoding topoisomerase II β binding protein 1 (TopBP1) exhibits severe definitive hematopoiesis failure. Homozygous topbp1cas003 mutants manifest reduced number of HSPCs during definitive hematopoietic cell expansion, without affecting the formation and migration of HSPCs. Moreover, HSPCs in the caudal hematopoietic tissue (an equivalent of the fetal liver in mammals) in topbp1cas003 mutant embryos are more sensitive to hydroxyurea (HU) treatment. Mechanistically, subcellular mislocalization of TopBP1cas003 protein results in ATR/Chk1 activation failure and DNA damage accumulation in HSPCs, and eventually induces the p53-dependent apoptosis of HSPCs. Collectively, this study demonstrates a novel and vital role of TopBP1 in the maintenance of HSPCs genome integrity and survival during hematopoietic progenitor expansion.

No MeSH data available.


Related in: MedlinePlus

The definitive hematopoiesis is defective in zebrafish mutantcas003 embryos.(A, A’) The bright field images of zebrafish wild-type sibling (A) and mutantcas003 embryos (A’) showing no obvious difference at 5dpf. (B-G’) Whole-mount in situ hybridization (WISH) results of c-myb, ae1-globin, mpx, lyz and rag1 showing defective definitive hematopoiesis in mutantcas003 embryos (B’-G’) but not in sibling embryos (B-G) at 5dpf. The penetrance of the indicated phenotype is shown in the bottom left of each panel. (B-F, B’-F’) Lateral views; (G, G’) dorsal views. Black arrows indicate the position of caudal hematopoietic tissue (CHT); red arrows and circles show the position of thymus and kidney, respectively.
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pgen.1005346.g001: The definitive hematopoiesis is defective in zebrafish mutantcas003 embryos.(A, A’) The bright field images of zebrafish wild-type sibling (A) and mutantcas003 embryos (A’) showing no obvious difference at 5dpf. (B-G’) Whole-mount in situ hybridization (WISH) results of c-myb, ae1-globin, mpx, lyz and rag1 showing defective definitive hematopoiesis in mutantcas003 embryos (B’-G’) but not in sibling embryos (B-G) at 5dpf. The penetrance of the indicated phenotype is shown in the bottom left of each panel. (B-F, B’-F’) Lateral views; (G, G’) dorsal views. Black arrows indicate the position of caudal hematopoietic tissue (CHT); red arrows and circles show the position of thymus and kidney, respectively.

Mentions: To explore new genes and regulatory mechanisms in vertebrate definitive hematopoiesis, we carried out a large-scale forward genetics screen on ENU-mutagenized F2 families in zebrafish by whole mount in situ hybridization (WISH) using c-myb probe (a key transcription factor and marker of HSPCs) [15,47]. In 5dpf wild-type zebrafish embryos, c-myb was expressed in all hematopoietic tissues including caudal hematopoietic tissue (CHT), thymus, and kidney (Fig 1); whereas homozygous mutantscas003 displayed normal morphogenesis (Fig 1A–A’), but dramatically decreased c-myb expression in CHT, kidney and thymus (Fig 1B–B’), suggesting the expansion of HSPCs was defective. To confirm the defective definitive hematopoiesis in mutantscas003, we further examined the expression of downstream hematopoietic lineage cell markers including ae1-globin (erythrocyte marker), mpx (granulocyte marker), lyz (macrophage marker) and rag1 (lymphocyte marker). The expression of all these markers was substantially decreased in the homozygous mutantcas003 embryos at 5dpf (Fig 1C–G’), which suggested hematopoiesis failure.


TopBP1 Governs Hematopoietic Stem/Progenitor Cells Survival in Zebrafish Definitive Hematopoiesis.

Gao L, Li D, Ma K, Zhang W, Xu T, Fu C, Jing C, Jia X, Wu S, Sun X, Dong M, Deng M, Chen Y, Zhu W, Peng J, Wan F, Zhou Y, Zon LI, Pan W - PLoS Genet. (2015)

The definitive hematopoiesis is defective in zebrafish mutantcas003 embryos.(A, A’) The bright field images of zebrafish wild-type sibling (A) and mutantcas003 embryos (A’) showing no obvious difference at 5dpf. (B-G’) Whole-mount in situ hybridization (WISH) results of c-myb, ae1-globin, mpx, lyz and rag1 showing defective definitive hematopoiesis in mutantcas003 embryos (B’-G’) but not in sibling embryos (B-G) at 5dpf. The penetrance of the indicated phenotype is shown in the bottom left of each panel. (B-F, B’-F’) Lateral views; (G, G’) dorsal views. Black arrows indicate the position of caudal hematopoietic tissue (CHT); red arrows and circles show the position of thymus and kidney, respectively.
© Copyright Policy
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4488437&req=5

pgen.1005346.g001: The definitive hematopoiesis is defective in zebrafish mutantcas003 embryos.(A, A’) The bright field images of zebrafish wild-type sibling (A) and mutantcas003 embryos (A’) showing no obvious difference at 5dpf. (B-G’) Whole-mount in situ hybridization (WISH) results of c-myb, ae1-globin, mpx, lyz and rag1 showing defective definitive hematopoiesis in mutantcas003 embryos (B’-G’) but not in sibling embryos (B-G) at 5dpf. The penetrance of the indicated phenotype is shown in the bottom left of each panel. (B-F, B’-F’) Lateral views; (G, G’) dorsal views. Black arrows indicate the position of caudal hematopoietic tissue (CHT); red arrows and circles show the position of thymus and kidney, respectively.
Mentions: To explore new genes and regulatory mechanisms in vertebrate definitive hematopoiesis, we carried out a large-scale forward genetics screen on ENU-mutagenized F2 families in zebrafish by whole mount in situ hybridization (WISH) using c-myb probe (a key transcription factor and marker of HSPCs) [15,47]. In 5dpf wild-type zebrafish embryos, c-myb was expressed in all hematopoietic tissues including caudal hematopoietic tissue (CHT), thymus, and kidney (Fig 1); whereas homozygous mutantscas003 displayed normal morphogenesis (Fig 1A–A’), but dramatically decreased c-myb expression in CHT, kidney and thymus (Fig 1B–B’), suggesting the expansion of HSPCs was defective. To confirm the defective definitive hematopoiesis in mutantscas003, we further examined the expression of downstream hematopoietic lineage cell markers including ae1-globin (erythrocyte marker), mpx (granulocyte marker), lyz (macrophage marker) and rag1 (lymphocyte marker). The expression of all these markers was substantially decreased in the homozygous mutantcas003 embryos at 5dpf (Fig 1C–G’), which suggested hematopoiesis failure.

Bottom Line: Homozygous topbp1cas003 mutants manifest reduced number of HSPCs during definitive hematopoietic cell expansion, without affecting the formation and migration of HSPCs.Moreover, HSPCs in the caudal hematopoietic tissue (an equivalent of the fetal liver in mammals) in topbp1cas003 mutant embryos are more sensitive to hydroxyurea (HU) treatment.Collectively, this study demonstrates a novel and vital role of TopBP1 in the maintenance of HSPCs genome integrity and survival during hematopoietic progenitor expansion.

View Article: PubMed Central - PubMed

Affiliation: Key Laboratory of Stem Cell Biology, Institute of Health Sciences, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences & Shanghai Jiao Tong University School of Medicine, Shanghai, China.

ABSTRACT
In vertebrate definitive hematopoiesis, nascent hematopoietic stem/progenitor cells (HSPCs) migrate to and reside in proliferative hematopoietic microenvironment for transitory expansion. In this process, well-established DNA damage response pathways are vital to resolve the replication stress, which is deleterious for genome stability and cell survival. However, the detailed mechanism on the response and repair of the replication stress-induced DNA damage during hematopoietic progenitor expansion remains elusive. Here we report that a novel zebrafish mutantcas003 with nonsense mutation in topbp1 gene encoding topoisomerase II β binding protein 1 (TopBP1) exhibits severe definitive hematopoiesis failure. Homozygous topbp1cas003 mutants manifest reduced number of HSPCs during definitive hematopoietic cell expansion, without affecting the formation and migration of HSPCs. Moreover, HSPCs in the caudal hematopoietic tissue (an equivalent of the fetal liver in mammals) in topbp1cas003 mutant embryos are more sensitive to hydroxyurea (HU) treatment. Mechanistically, subcellular mislocalization of TopBP1cas003 protein results in ATR/Chk1 activation failure and DNA damage accumulation in HSPCs, and eventually induces the p53-dependent apoptosis of HSPCs. Collectively, this study demonstrates a novel and vital role of TopBP1 in the maintenance of HSPCs genome integrity and survival during hematopoietic progenitor expansion.

No MeSH data available.


Related in: MedlinePlus