Regulation of mouse lens fiber cell development and differentiation by the Maf gene

Maf is a basic domain/leucine zipper domain protein originally identified as a proto-oncogene whose consensus target site in vitro, the T-MARE, is an extended version of an AP-1 site normally recognized by Fos and Jun. Maf and the closely related family members Neural retina leucine zipper (Nrl), L-Maf, and Krml1/MafB have been implicated in a wide variety of developmental and physiologic roles; however, mutations in vivo have been described only for Krml1/MafB, in which a loss-of-function causes abnormalities in hindbrain development due to failure to activate the Hoxa3 and Hoxb3 genes. We have used gene targeting to replace Maf coding sequences with those of lacZ, and have carried out a comprehensive analysis of embryonic expression and the homozygous mutant phenotype in the eye. Maf is expressed in the lens vesicle after invagination, and becomes highly upregulated in the equatorial zone, the site at which self-renewing anterior epithelial cells withdraw from the cell cycle and terminally differentiate into posterior fiber cells. Posterior lens cells in Maf(lacZ) mutant mice exhibit failure of elongation at embryonic day 11.5, do not express (&agr;)A- and all of the (beta)-crystallin genes, and display inappropriately high levels of DNA synthesis. This phenotype partially overlaps with those reported for gene targeting of Prox1 and Sox1; however, expression of these genes is grossly normal, as is expression of Eya1, Eya2, Pax6, and Sox2. Recombinant Maf protein binds to T-MARE sites in the (alpha)A-, (beta)B2-, and (beta)A4-crystallin promoters but fails to bind to a point mutation in the (alpha)A-crystallin promoter that has been shown previously to be required for promoter function. Our results indicate that Maf directly activates many if not all of the (beta)-crystallin genes, and suggest a model for coordinating cell cycle withdrawal with terminal differentiation.     

Structure-function analysis of the maf oncogene product, a member of the b-Zip protein family.

The v-maf oncogene, identified as the transforming gene of the avian retrovirus AS42, encodes a protein containing a b-Zip motif. From this structural feature, the v-Maf protein was expected to form a dimer and function as a nuclear DNA-binding protein. In this study, we demonstrate that this protein indeed localizes predominantly in the nucleus and forms a homodimer through its leucine zipper structure. To delineate the structural requirement for the transforming activity, we constructed and characterized a panel of v-maf mutants harboring various deletions or point mutations. A region of about 100 amino acid residues located near its carboxyl terminus, which contains the b-Zip motif, was found to be essential for the basal transforming activity of v-Maf on chicken embryo fibroblasts. On the other hand, the amino-terminal two-thirds of the v-Maf protein seems to play a role in potentiating the transforming activity of v-Maf. It was also found that the c-maf proto-oncogene, without any structural modification in its protein-coding region, could transform cells as efficiently as could the v-maf oncogene when transduced by a retroviral vector. Thus, it is probably deregulated expression that makes the v-maf gene oncogenic. In addition, we discovered one point mutation, altering the structure of the b-Zip domain, which further enhances the transforming activity of the v-maf oncogene. Such mutant will be useful in exploring the mechanism of action of the Maf protein.     

胰腺发育相关maf基因在胰腺导管和胰岛的表达

为探讨胰岛功能和发育相关maf基因在胰腺导管上皮中的表达情况,对新鲜小鼠胰腺组织切片进行显微切割,分离纯化胰腺组织中的导管和胰岛,以及外分泌腺组织细胞作为对照,利用荧光实时定量PCR的方法完成对目的基因的相对定量.结果显示,mafa mRNA,mafb mRNA水平在胰岛及导管中非常接近,无统计学差异.而c-maf在导管的表达高于胰岛(P<0.05),外分泌腺则无上述基因的表达.胰腺导管中mafa,mafb,cmaf均有表达,肯定了导管上皮细胞向内分泌细胞分化的潜能,而c-maf在导管中的表达高于胰岛,提示导管上皮c-maf的下调可能有助于导管上皮细胞向内分泌细胞的分化成熟.