Institute of Plant and Microbial Biology, Academia Sinica, Taipei, Taiwan 11529.
Plant Physiol. 2013 Nov;163(3):1218-29. doi: 10.1104/pp.113.225706. Epub 2013 Oct 4.
Lipid transfer proteins (LTPs) are small secretory proteins in plants with defined lipid-binding structures for possible lipid exocytosis. Special groups of LTPs unique to the anther tapetum are abundant, but their functions are unclear. We studied a special group of LTPs, type III LTPs, in Arabidopsis (Arabidopsis thaliana). Their transcripts were restricted to the anther tapetum, with levels peaking at the developmental stage of maximal pollen-wall exine synthesis. We constructed an LTP-Green Fluorescent Protein (LTP-GFP) plasmid, transformed it into wild-type plants, and monitored LTP-GFP in developing anthers with confocal laser scanning microscopy. LTP-GFP appeared in the tapetum and was secreted via the endoplasmic reticulum-trans-Golgi network machinery into the locule. It then moved to the microspore surface and remained as a component of exine. Immuno-transmission electron microscopy of native LTP in anthers confirmed the LTP-GFP observations. The in vivo association of LTP-GFP and exine in anthers was not observed with non-type III or structurally modified type III LTPs or in transformed exine-defective mutant plants. RNA interference knockdown of individual type III LTPs produced no observable mutant phenotypes. RNA interference knockdown of two type III LTPs produced microscopy-observable morphologic changes in the intine underneath the exine (presumably as a consequence of changes in the exine not observed by transmission electron microscopy) and pollen susceptible to dehydration damage. Overall, we reveal a novel transfer pathway of LTPs in which LTPs bound or nonbound to exine precursors are secreted from the tapetum to become microspore exine constituents; this pathway explains the need for plentiful LTPs to incorporate into the abundant exine.
脂质转移蛋白（LTPs）是植物中具有特定脂质结合结构的小分泌蛋白，可能参与脂质胞吐作用。花药绒毡层中存在大量特有的特殊 LTP 蛋白，但它们的功能尚不清楚。我们研究了拟南芥（Arabidopsis thaliana）中一种特殊的 LTP 蛋白，即 III 型 LTP 蛋白。它们的转录本仅限于花药绒毡层，在花粉外壁 exine 合成的最旺盛的发育阶段达到峰值。我们构建了一个 LTP-绿色荧光蛋白（LTP-GFP）质粒，将其转化为野生型植物，并通过共焦激光扫描显微镜监测发育中的花药中的 LTP-GFP。LTP-GFP 出现在绒毡层中，并通过内质网-高尔基体网络机制被分泌到腔室中。然后，它移动到花粉表面，并作为 exine 的组成部分保留下来。对花药中天然 LTP 的免疫透射电镜观察证实了 LTP-GFP 的观察结果。在非 III 型或结构修饰的 III 型 LTPs 或转化的 exine 缺陷突变体植物中，未观察到 LTP-GFP 与 exine 的体内关联。单个 III 型 LTP 的 RNA 干扰敲低未产生可观察到的突变表型。两个 III 型 LTP 的 RNA 干扰敲低导致 exine 下的内体（推测是由于透射电镜未观察到的 exine 变化）产生显微镜可观察的形态变化，以及对脱水损伤敏感的花粉。总的来说，我们揭示了一种新型的 LTP 转移途径，其中与 exine 前体结合或未结合的 LTP 从绒毡层分泌出来，成为花粉外壁 exine 的组成部分；这种途径解释了需要大量的 LTP 掺入到丰富的 exine 中。
