Henry Kelli F, Kawashima Tomokazu, Goldberg Robert B
Department of Molecular, Cell and Developmental Biology, University of California, Los Angeles, 610 Charles E. Young Dr. East, Los Angeles, CA, 90095-7239, USA.
Plant Mol Biol. 2015 Jun;88(3):207-17. doi: 10.1007/s11103-015-0308-z. Epub 2015 Mar 22.
Little is known about the molecular mechanisms by which the embryo proper and suspensor of plant embryos activate specific gene sets shortly after fertilization. We analyzed the upstream region of the Scarlet Runner Bean (Phaseolus coccineus) G564 gene in order to understand how genes are activated specifically in the suspensor during early embryo development. Previously, we showed that a 54-bp fragment of the G564 upstream region is sufficient for suspensor transcription and contains at least three required cis-regulatory sequences, including the 10-bp motif (5'-GAAAAGCGAA-3'), the 10 bp-like motif (5'-GAAAAACGAA-3'), and Region 2 motif (partial sequence 5'-TTGGT-3'). Here, we use site-directed mutagenesis experiments in transgenic tobacco globular-stage embryos to identify two additional cis-regulatory elements within the 54-bp cis-regulatory module that are required for G564 suspensor transcription: the Fifth motif (5'-GAGTTA-3') and a third 10-bp-related sequence (5'-GAAAACCACA-3'). Further deletion of the 54-bp fragment revealed that a 47-bp fragment containing the five motifs (the 10-bp, 10-bp-like, 10-bp-related, Region 2 and Fifth motifs) is sufficient for suspensor transcription, and represents a cis-regulatory module. A consensus sequence for each type of motif was determined by comparing motif sequences shown to activate suspensor transcription. Phylogenetic analyses suggest that the regulation of G564 is evolutionarily conserved. A homologous cis-regulatory module was found upstream of the G564 ortholog in the Common Bean (Phaseolus vulgaris), indicating that the regulation of G564 is evolutionarily conserved in closely related bean species.
关于植物胚胎的胚体和胚柄在受精后不久激活特定基因集的分子机制，我们了解得还很少。我们分析了红花菜豆（Phaseolus coccineus）G564基因的上游区域，以了解在胚胎早期发育过程中基因是如何在胚柄中特异性激活的。此前，我们表明G564上游区域的一个54 bp片段足以进行胚柄转录，并且包含至少三个必需的顺式调控序列，包括10 bp基序（5'-GAAAAGCGAA-3'）、10 bp样基序（5'-GAAAAACGAA-3'）和区域2基序（部分序列5'-TTGGT-3'）。在这里，我们在转基因烟草球形期胚胎中进行定点诱变实验，以鉴定54 bp顺式调控模块内另外两个G564胚柄转录所需的顺式调控元件：第五基序（5'-GAGTTA-3'）和第三个10 bp相关序列（5'-GAAAACCACA-3'）。对54 bp片段的进一步缺失分析表明，一个包含五个基序（10 bp、10 bp样、10 bp相关、区域2和第五基序）的47 bp片段足以进行胚柄转录，并代表一个顺式调控模块。通过比较显示可激活胚柄转录的基序序列，确定了每种基序类型的共有序列。系统发育分析表明，G564的调控在进化上是保守的。在菜豆（Phaseolus vulgaris）中G564直系同源基因的上游发现了一个同源顺式调控模块，这表明G564的调控在近缘豆科物种中在进化上是保守的。
