Controlling pluripotency and differentiation in plant cells: the KNOX-TCP regulatory module

多细胞生物的发育涉及到从祖干细胞群体中分化出特定的细胞、组织和器官类型。在拟南芥（Arabidopsis thaliana）等植物中，多能干细胞位于未分化的细胞池中，称为茎尖分生组织（shoot apical meristem， SAM），该细胞产生侧面器官，如叶片。拟南芥是植物发育生物学研究中常用的模式生物。随后，叶细胞经历与它们的特殊功能相关的分化，如光合作用和气体交换。已经确定了许多基因参与控制SAM的形成和维持功能。一些基因的作用是赋予多能状态——也就是说，一种能够形成任何类型细胞的未分化状态——而另一些基因则促进分化成专门的细胞类型。目前已知的大多数调节SAM功能的基因编码一种称为转录因子（TF）的蛋白质。tf是一种蛋白质，它通过激活或抑制其他所谓的“靶”基因的表达来调节它们的表达，特别是通过控制基因从DNA到RNA的转录，RNA随后被翻译成功能性蛋白质。在已被证明参与SAM调控的许多不同类型的TF中，knoted1样同源盒（KNOX）基因SHOOT MERISTEMLESS （STM）编码的TF对SAM中多能细胞命运的规范和维持绝对至关重要。我最近的研究表明，STM抑制了TCP家族（TCP4）中另一种TF的表达，该基因参与促进叶片分化。有趣的是，TCP tf先前已被证明可以抑制STM的表达。因此，我揭示了这两类重要的TF之间的相互拮抗关系：STM促进SAM的多能性并抑制TCP的表达，从而阻止分生组织细胞的分化，而TCP抑制STM的表达并促进发育叶片的分化。STM功能丧失或TCP水平升高会导致SAM细胞分化并停止活动。相反，TCP的缺失模拟了STM表达升高的影响，如抑制叶片分化。鉴于这些相似之处，人们可能会期望STM和tcp以拮抗的方式调节一组共同的靶基因，这些基因导致细胞采用多能或分化的命运。多能性和分化的控制是本研究项目的中心主题。具体来说，该项目旨在通过研究STM和TCP之间的相互作用，确定TCP调控的靶基因，并将这些基因与先前确定的STM基因进行比较，来了解SAM中细胞命运决定是如何做出的。利用这两个基因的荧光“报告者”，当TCP或STM水平分别被人为地增加或减少时，将监测活体SAM中STM和TCP表达的变化，揭示它们在哪些细胞中表达变化以及这种变化发生的速度。在先前确定了促进多能性的STM靶基因之后，对TCP靶基因的表征将使鉴定促进分化的靶基因成为可能。比较分析将揭示哪些基因受到STM和TCP的竞争性调控。这些可能代表了多能性或分化细胞命运的关键参与者，并代表了我们对植物发育中多能性和分化控制的理解取得了相当大的进步。

Development in multicellular organisms involves the differentiation of specific cell-, tissue- and organ-types from progenitor stem cell populations. In plants such as Arabidopsis thaliana, a commonly used model organism used in the study of plant developmental biology, pluripotent stem cells are located in a pool of undifferentiated cells termed the shoot apical meristem (SAM), which gives rise to lateral organs such as leaves. Leaf cells subsequently undergo differentiation associated with their specialised functions, such as photosynthesis and gas exchange. Numerous genes have been identified that are involved in controlling the formation and sustained function of the SAM. Some genes act to confer a state of pluripotency - that is to say, an undifferentiated state with the ability to form any type of cell - while others promote differentiation into specialised cell types. Most of the genes currently known to regulate SAM function encode a type of protein called a transcription factor (TF). TFs are proteins that regulate the expression of other so-called 'target' genes by activating or repressing their expression - specifically by controlling the transcription of a gene from DNA into RNA, which is subsequently translated into a functional protein. Of the many different types of TF that have been shown to be involved in SAM regulation, the KNOTTED1-like homeobox (KNOX) gene SHOOT MERISTEMLESS (STM) encodes a TF that is absolutely critical for the specification and maintenance of pluripotent cell fate in the SAM. My recent work has shown that STM represses the expression of a different type of TF belonging to the TCP family (TCP4), which is involved in promoting differentiation in leaves. Interestingly, TCP TFs have previously been shown to repress the expression of STM. Hence, I have revealed a mutually antagonistic relationship between these two important classes of TF: STM promotes pluripotency and represses TCP expression in the SAM, thereby preventing meristem cell differentiation, while TCP represses STM expression and promotes differentiation in developing leaves. Loss of STM function or elevated levels of TCP causes the cells of the SAM to differentiate and cease activity. Conversely, loss of TCP mimics the effect of elevated STM expression, such as inhibited leaf differentiation. Given these similarities, one might expect that STM and TCPs regulate, in an antagonistic manner, a common set of target genes that lead to cells adopting pluripotent or differentiated fates.The control of pluripotency and differentiation is the central theme of this research project. Specifically, this project seeks to understand how cell fate decisions are made in the SAM by studying the interplay between STM and TCP, identifying TCP-regulated target genes and comparing these with those previously identified for STM. Using fluorescent 'reporters' for these two genes, changes in STM and TCP expression in the living SAM will be monitored when levels of TCP or STM levels are artificially increased or decreased respectively, revealing in which cells their expression changes and the how quickly this occurs. Having previously identified the STM target genes that promote pluripotency, characterisation of TCP target genes will enable the identification of target genes that promote differentiation. Comparative analyses will reveal which genes are competitively regulated by STM and TCP. These would likely represent key players for the adoption of pluripotent or differentiated cell fates and would represent a considerable advance in our understanding of the control of pluripotency and differentiation in plant development.