School of Life Sciences, University of Dundee, Dow St., Dundee, DD1 5EH, UK.
Dev Biol. 2011 May 15;353(2):290-301. doi: 10.1016/j.ydbio.2011.03.003. Epub 2011 Mar 21.
Dictyostelium is the only non-metazoan with functionally analyzed SH2 domains and studying them can give insights into their evolution and wider potential. LrrB has a novel domain configuration with leucine-rich repeat, 14-3-3 and SH2 protein-protein interaction modules. It is required for the correct expression of several specific genes in early development and here we characterize its role in later, multicellular development. During development in the light, slug formation in LrrB null (lrrB-) mutants is delayed relative to the parental strain, and the slugs are highly defective in phototaxis and thermotaxis. In the dark the mutant arrests development as an elongated mound, in a hitherto unreported process we term dark stalling. The developmental and phototaxis defects are cell autonomous and marker analysis shows that the pstO prestalk sub-region of the slug is aberrant in the lrrB- mutant. Expression profiling, by parallel micro-array and deep RNA sequence analyses, reveals many other alterations in prestalk-specific gene expression in lrrB- slugs, including reduced expression of the ecmB gene and elevated expression of ampA. During culmination ampA is ectopically expressed in the stalk, there is no expression of ampA and ecmB in the lower cup and the mutant fruiting bodies lack a basal disc. The basal disc cup derives from the pstB cells and this population is greatly reduced in the lrrB- mutant. This anatomical feature is a hallmark of mutants aberrant in signaling by DIF-1, the polyketide that induces prestalk and stalk cell differentiation. In a DIF-1 induction assay the lrrB- mutant is profoundly defective in ecmB activation but only marginally defective in ecmA induction. Thus the mutation partially uncouples these two inductive events. In early development LrrB interacts physically and functionally with CldA, another SH2 domain containing protein. However, the CldA null mutant does not phenocopy the lrrB- in its aberrant multicellular development or phototaxis defect, implying that the early and late functions of LrrB are affected in different ways. These observations, coupled with its domain structure, suggest that LrrB is an SH2 adaptor protein active in diverse developmental signaling pathways.
盘基网柄菌是唯一具有功能分析的 SH2 结构域的非后生动物，研究它可以深入了解其进化和更广泛的潜力。LrrB 具有新颖的结构域结构，具有富含亮氨酸的重复序列、14-3-3 和 SH2 蛋白-蛋白相互作用模块。它是早期发育中几个特定基因正确表达所必需的，在这里我们描述了它在后期多细胞发育中的作用。在光下发育时，LrrB 缺失（lrrB-）突变体中的变形虫形成相对于亲本菌株延迟，并且变形虫在光和热趋性方面存在严重缺陷。在黑暗中，突变体作为伸长的土丘发育停滞，在一个我们称之为黑暗停滞的前所未有的过程中。发育和光趋性缺陷是细胞自主的，标记分析表明，lrrB-突变体中的变形虫的 pstO 前胃亚区是异常的。通过平行的微阵列和深度 RNA 序列分析进行表达谱分析，揭示了 lrrB-变形虫中前胃特异性基因表达的许多其他改变，包括 ecmB 基因的表达降低和 ampA 的表达升高。在 culmination 期间，ampA 在茎中异位表达，在下部杯中没有 ampA 和 ecmB 的表达，突变体的果体缺乏基部盘。基部盘杯源自 pstB 细胞，该细胞在 lrrB-突变体中大大减少。这个解剖特征是突变体中由 DIF-1 诱导的信号传导异常的标志，DIF-1 是诱导前胃和茎细胞分化的聚酮。在 DIF-1 诱导测定中，lrrB-突变体在 ecmB 的激活中严重缺陷，但在 ecmA 的诱导中仅轻微缺陷。因此，该突变体部分地使这两个诱导事件解偶联。在早期发育中，LrrB 与另一种含有 SH2 结构域的蛋白质 CldA 物理和功能相互作用。然而，CldA 缺失突变体在其异常的多细胞发育或光趋性缺陷中并不模拟 lrrB-，这意味着 LrrB 的早期和晚期功能以不同的方式受到影响。这些观察结果，加上其结构域结构，表明 LrrB 是一种在不同发育信号通路中起作用的 SH2 衔接蛋白。
