Gravitropic setpoint angle control in higher plants

植物的整体形状，即它们在地上和地下所占的空间，主要是由枝干的数量、长度和角度决定的。有趣的是，许多侧枝生长的角度是相对于重力而不是主要的根-枝轴来设定和维持的。这些角被称为地向设定角或gsa。尽管GSA是自然界中观察到的植物结构奇妙变化的基本组成部分，但直到最近，GSA调节的机制还不为人所知。主主根-茎轴垂直gsa的维持，也被称为向地性，是很容易理解的：主茎或根的位移是由根和芽中专门的重力感应细胞感知的。这导致一种叫做生长素的植物激素从这些细胞主动运输到移位器官的下部。在根中，生长素抑制细胞伸长，导致向下弯曲到垂直，而在茎中，生长素则相反，促进向上弯曲。在我们之前关于GSA控制的研究中，我们发现在非垂直的根和枝上也存在同样的向地性响应，但会被另一个生长成分——反向地性抵消抵消。我们发现，垂直度较低的分支具有更强的反地向偏移，并且这种偏移的大小也直接在重力感应细胞内调节。我们还表明，像向地向性一样，反向地向性也依赖于生长素的运输，但在这种情况下，是从重力感应细胞的上侧而不是下侧运输。反向地性活性的分子机制以及这种活性如何局限于非垂直分支尚不清楚，这是本研究的主题。这个项目有三个部分。首先，我们关注生长素转运体PIN蛋白的调控，PIN蛋白负责将生长素移出细胞。在重力感应细胞中，两种特殊的pin， PIN3和PIN7，似乎同时介导向地性和反向地性生长素通量。pin的亚细胞定位和活性已被证明是由它们的磷酸化调节的。我们的初步工作表明，突变的磷酸化版本PIN3具有较少的垂直侧根，而不能磷酸化的突变PIN3具有更多的垂直侧根。对这些数据最简单的解释是，重力感应细胞中未磷酸化的PIN有助于向下的、向地向的生长素通量，而磷酸化的PIN介导向上的、反向地向的通量。重要的是，虽然重力感应细胞的向地性通量方向可以在植物重新定向时非常迅速地重置，但我们的数据表明，驱动反向地性活动的生长素通量方向的变化需要更长的时间才能重新建立。因此，通过跟踪侧根重力感应细胞中发生简单定向的分子和细胞生物学事件，我们可以观察到哪些PIN和PIN磷调节因子与曾经运输生长素以维持向地性或反向地性生长的细胞的表面相关，然后观察到向地性和反向地性生长素通量的新极性被重新建立。向重力性的一个主要特征是，一个器官离垂直方向越远，重力反应的强度就越大。这种现象对GSA的控制至关重要，因此在项目的第二部分，我们使用先进的显微镜来了解这种反应的生物物理和分子基础。在项目的最后一部分，我们将筛选一组突变的拟南芥植株，寻找侧枝很少或没有非垂直生长的植株。这将使我们能够识别限制抗向地性活性的基因，并使我们能够形成对植物中GSA控制的连贯理解。

The overall shape of plants, the space they occupy above and below ground, is determined principally by the number, length, and angle of their branches. Interestingly, the angles at which many lateral branches grow out are set and maintained relative to gravity rather than the main root-shoot axis. These angles are known as a gravitropic setpoint angles or GSAs. Despite being a fundamental component of the wonderful variation in plant architecture observed throughout nature until recently the mechanisms underlying GSA regulation were not known. The maintenance of vertical GSAs in the main primary root-shoot axis, otherwise known as gravitropism, is well understood: displacement of the main stem or root is perceived within specialised gravity-sensing cells in roots and shoots. This leads to the active transport of a plant hormone called auxin from these cells and to the lower side of the displaced organ. In roots, auxin inhibits cell elongation, causing bending downwards to the vertical while in the shoot, auxin does the opposite, promoting upward curvature. In our previous work on GSA control we showed that the same underlying gravitropic response occurs in non-vertical root and shoot branches but is counteracted by another growth component, the antigravitropic offset. We showed that branches that are less vertical have a stronger antigravitropic offset and that the magnitude of this offset is also regulated directly within the gravity-sensing cell. We have also shown that like gravitropism, antigravitropism depends on auxin transport but in this case from the upper, rather than the lower side of the gravity-sensing cell. The molecular mechanisms underlying antigravitropic activity and how that activity is restricted to non-vertical branches is not known and is the subject of this study.The project has three parts. In the first, we focus on the regulation auxin transporters called PIN proteins that are responsible for moving auxin out of cells. In gravity-sensing cells, two particular PINs, PIN3 and PIN7, appear to mediate both gravitropic and antigravitropic auxin fluxes. The subcellular localisation and activity of PINs has been shown to be regulated by their phosphorylation. Our preliminary work has shown that a mutated phosphomimic version of PIN3 has less vertical lateral roots while a mutated PIN3 that cannot be phosphorylated has more vertical lateral roots. The simplest interpretation of these data is that unphosphorylated PINs in gravity-sensing cells contribute to downward, gravitropic auxin flux while phosphorylated PIN mediates an upward, antigravitropic flux. Importantly, while the direction of the gravitropic flux out of the gravity-sensing cells can be reset very rapidly if the plant is reorientated, our data indicate that changes in the direction of auxin fluxes that drive antigravitropic activity take much longer to be re-established. Therefore, by tracking the molecular and cell biological events within gravity-sensing cells of lateral roots undergoing simple reorientations we can observe which PINs and PIN phosphoregulators are associated with the faces of the cell that were once transporting auxin to sustain gravitropic or antigravitropic growth and then watch as the new polarities for gravi- and antigravitropic auxin fluxes are re-established.A central feature of gravitropism is that the magnitude of graviresponse increases the further an organ is moved away from the vertical. This phenomenon is crucial for GSA control and so in the second part of the project we use advanced microscopy to understand the biophysical and molecular basis of this response. In the final part of the project we will screen a collection of mutated Arabidopsis plants to look for plants with little or no non-vertical growth in lateral branches. This will allow us to identify the genes that restrict antigravitropic activity to lateral branches and allow us to form a coherent understanding of GSA control in plants.