丝裂原活化蛋白激酶磷酸酶-2(MKP-2/DUSP4)具有酪氨酸磷酸酶和丝氨酸/苏氨酸磷酸酶活性, 可以作用于MAPKs(Mitogen-activated protein kinases)底物, 使其去磷酸化。但dusp4在脊椎动物胚胎发育中的功能还所知甚少。为深入了解dusp4在发育中的作用, 文章首先检测了其在斑马鱼胚胎的表达。通过整胚原位杂交实验, 发现dusp4是斑马鱼母源表达的基因, 并且随着发育的进行, 在原肠早期特异表达在中内胚层区域。进一步的实验表明, Nodal信号对dusp4的表达至关重要。过表达Nodal信号的配体sqt、dusp4的表达水平明显升高, 而在缺失Nodal信号的突变体MZoep中, 几乎检测不到dusp4的表达。此外, 文章利用反义核苷酸Morpholino (MO)敲降dusp4的表达, 中内胚层标识基因gsc、sox17和sox32的表达水平显著升高, 而过表达dusp4对中内胚层的形成没有明显的影响, 表明dusp4对中内胚层形成的抑制作用是必需的, 但不是充分的, 可能还有其他未被鉴定的协同作用因子。以上研究结果表明, dusp4基因的表达受到Nodal信号调控, 在原肠期具有抑制中内胚层形成的作用。
MAP kinase phosphatase-2 (MKP-2/DUSP4), a dual specificity protein phosphatase with tyrosine/serine/ threonine phosphatase activity, is associated with cellular proliferation and differentiation, but its functions during embryo development are unclear. To study the developmental function of dusp4, we first examined the spatiotemporal ex-pression pattern of this gene during zebrafish embryonic development by whole mount in situ hybridization. We found that dusp4 was maternally expressed since its transcripts were present from the one-cell to the 256-cell stages. At early gastrulation stages, dusp4 transcripts specifically distributed at margin region, where the mesendodermal cells were located. Furthermore, Nodal signal was crucial for dusp4 expression. The expression of dusp4 was obviously increased in Nodal ligand overexpressed embryos, while its expression was almost lost in the Nodal sig-nal-deficient MZoep mutants. In addition, dusp4 MO was also designed to knock down its expression in embryos. The mesendoderm formation was significantly increased in dusp4 morphants, but not obviously changed in dusp4 overexpressed embryos, suggesting that dusp4 is necessary, but not sufficient for the inhibitory of mesendoderm induction. Thus, our results indicate that Nodal regulated dusp4 plays a repressive role in mesendoderm induction.
[1] Schier AF. Nodal signaling in vertebrate development. Annu Rev Cell Dev Biol, 2003, 19: 589-621.
[2] Whitman M. Nodal signaling in early vertebrate embryos: themes and variations. Dev Cell, 2001, 1(5): 605-617.
[3] Tian T, Meng AM. Nodal signals pattern vertebrate em-bryos. Cell Mol Life Sci, 2006, 63(6): 672-685.
[4] Shen MM, Schier AF. The EGF-CFC gene family in vertebrate development. Trends Genet, 2000, 16(7): 303-309.
[5] Zhang JJ, Talbot WS, Schier AF. Positional cloning iden-tifies zebrafish one-eyed pinhead as a permissive EGF-related ligand required during gastrulation. Cell, 1998, 92(2): 241-251.
[6] Gritsman K, Zhang JJ, Cheng S, Heckscher E, Talbot WS, Schier AF. The EGF-CFC protein one-eyed pinhead is es-sential for nodal signaling. Cell, 1999, 97(1): 121-132.
[7] Feldman B, Gates MA, Egan ES, Dougan ST, Rennebeck G, Sirotkin HI, Schier AF, Talbot WS. Zebrafish organizer development and germ-layer formation require nodal-related signals. Nature, 1998, 395(6698): 181-185.
[8] Massagué J. TGF-β signal transduction. Annu Rev Bio-chem, 1998, 67: 753-791.
[9] Massagué J, Seoane J, Wotton D. Smad transcription fac-tors. Genes Dev, 2005, 19(23): 2783-2810.
[10] Liu ZT, Lin XW, Cai ZP, Zhang ZQ, Han CS, Jia SJ, Meng AM, Wang Q. Global identification of SMAD2 target genes reveals a role for multiple co-regulatory factors in zebrafish early gastrulas. J Biol Chem, 2011, 286(32): 28520-28532.
[11] Patterson KI, Brummer T, O'Brien PM, Daly RJ. Dual-specificity phosphatases: critical regulators with diverse cellular targets. Biochem J, 2009, 418(3): 475-489.
[12] Nunes-Xavier C, Roma-Mateo C, Ríos P, Tárrega C, Ce-judo-Marín R, Tabernero L, Pulido R. Dual-specificity MAP kinase phosphatases as targets of cancer treatment. Anticancer Agents Med Chem, 2011, 11(1): 109-132.
[13] Brondello JM, Brunet A, Pouysségur J, McKenzie FR. The dual specificity mitogen-activated protein kinase phos-phatase-1 and -2 are induced by the p42/p44MAPK cas-cade. J Biol Chem, 1997, 272(2): 1368-1376.
[14] Chen PL, Hutter D, Yang XL, Gorospe M, Davis RJ, Liu YS. Discordance between the binding affinity of mitogen-activated protein kinase subfamily members for MAP kinase phosphatase-2 and their ability to activate the phos-phatase catalytically. J Biol Chem, 2001, 276(31): 29440-29449.
[15] Robinson CJM, Sloss CM, Plevin R. Inactivation of JNK activity by mitogen-activated protein kinase phos-phatase-2 in EAhy926 endothelial cells is dependent upon agonist-specific JNK translocation to the nucleus. Cell Signal, 2001, 13(1): 29-41.
[16] Jeffrey KL, Camps M, Rommel C, Mackay CR. Targeting dual-specificity phosphatases: manipulating MAP kinase signalling and immune responses. Nat Rev Drug Dis-cov, 2007, 6(5): 391-403.
[17] Ramesh S, Qi XJ, Wildey GM, Robinson J, Molkentin J, Letterio J, Howe PH. TGFβ-mediated BIM expression and apoptosis are regulated through SMAD3-dependent expression of the MAPK phosphatase MKP2. EMBO Rep, 2008, 9(10): 990-997.
[18] Brown JL, Snir M, Noushmehr H, Kirby M, Hong SK, Elkahloun AG, Feldman B. Transcriptional profiling of endogenous germ layer precursor cells identifies dusp4 as an essential gene in zebrafish endoderm specification. Proc Natl Acad Sci USA, 2008, 105(34): 12337-12342.
[19] Bennett JT, Joubin K, Cheng S, Aanstad P, Herwig R, Clark M, Lehrach H, Schier AF. Nodal signaling activates differentiation genes during zebrafish gastrulation. Dev Biol, 2007, 304(2): 525-540.
[20] Tseng WF, Jang TH, Huang CB, Yuh CH. An evolutionarily conserved kernel of gata5, gata6, otx2 and prdm1a operates in the formation of endoderm in zebrafish. Dev Biol, 2011, 357(2): 541-557.
[21] Ao
