Okamoto K, Perlman P S, Butow R A
Department of Molecular Biology and Oncology, University of Texas Southwestern Medical Center, Dallas, Texas 75235-9148, USA.
J Cell Biol. 1998 Aug 10;142(3):613-23. doi: 10.1083/jcb.142.3.613.
Green fluorescent protein (GFP) was used to tag proteins of the mitochondrial matrix, inner, and outer membranes to examine their sorting patterns relative to mtDNA in zygotes of synchronously mated yeast cells in rho+ x rho0 crosses. When transiently expressed in one of the haploid parents, each of the marker proteins distributes throughout the fused mitochondrial reticulum of the zygote before equilibration of mtDNA, although the membrane markers equilibrate slower than the matrix marker. A GFP-tagged form of Abf2p, a mtDNA binding protein required for faithful transmission of rho+ mtDNA in vegetatively growing cells, colocalizes with mtDNA in situ. In zygotes of a rho+ x rho+ cross, in which there is little mixing of parental mtDNAs, Abf2p-GFP prelabeled in one parent rapidly equilibrates to most or all of the mtDNA, showing that the mtDNA compartment is accessible to exchange of proteins. In rho+ x rho0 crosses, mtDNA is preferentially transmitted to the medial diploid bud, whereas mitochondrial GFP marker proteins distribute throughout the zygote and the bud. In zygotes lacking Abf2p, mtDNA sorting is delayed and preferential sorting is reduced. These findings argue for the existence of a segregation apparatus that directs mtDNA to the emerging bud.
绿色荧光蛋白(GFP)被用于标记线粒体基质、内膜和外膜的蛋白质,以研究在rho⁺×rho⁰杂交的同步交配酵母细胞合子中,它们相对于线粒体DNA(mtDNA)的分选模式。当在其中一个单倍体亲本中瞬时表达时,在mtDNA平衡之前,每个标记蛋白都分布在合子融合的线粒体网状结构中,尽管膜标记物的平衡比基质标记物慢。Abf2p是营养生长细胞中rho⁺ mtDNA忠实传递所需的一种mtDNA结合蛋白,其GFP标记形式在原位与mtDNA共定位。在rho⁺×rho⁺杂交的合子中,亲本mtDNA几乎没有混合,在一个亲本中预先标记的Abf2p-GFP迅速平衡到大部分或所有的mtDNA,表明mtDNA区室可进行蛋白质交换。在rho⁺×rho⁰杂交中,mtDNA优先传递到中间的二倍体芽,而线粒体GFP标记蛋白分布在整个合子和芽中。在缺乏Abf2p的合子中,mtDNA分选延迟,优先分选减少。这些发现支持存在一种将mtDNA导向新生芽的分离机制。