实验海洋生物学重点实验室知识库

Hox基因是动物发育过程的重要调控基因，在体轴决定中发挥重要功能。Hox基因的表达规律在不同动物中呈现很高的相似性，多沿前后轴顺序表达（空间共线性）。然而，在软体动物中没有发现这种规律。目前对软体动物Hox基因表达模式研究表明，不同动物的Hox基因表达模式非常多样化，即使在亲缘关系较近的类群中也能观察到差异很大的结果。部分研究发现了一些Hox基因存在共线性表达的证据，但是表现出共线性表达特征的组织却不一致。此外，即使不考虑共线性表达，软体动物的Hox表达仍然无法找到一个统一的规律。研究中经常发现Hox表达与幼虫贝壳形成区、足组织以及神经系统发育存在相关性，但是在这些器官中，Hox基因的表达模式似乎仍然是多种多样的。因此，软体动物Hox基因表达及功能是目前发育和演化研究中尚未解决的一个难题。

基于这种现状，我们推测，Hox的基因表达可能随发育的进行而存在较快速的变化，由此导致了在不同阶段观察到的表达规律不一致。为了验证这个假设，本研究选取了亲缘关系较远的两种软体动物：隶属于腹足纲的笠贝（Lottia goshimai）和隶属于多板纲的石鳖（Acanthochitona rubrolineata）。以这两种动物作为研究对象，研究了不同发育阶段Hox基因的表达模式，以期找到一种普适的Hox基因表达规律，为软体动物的发育和演化提供基础支持。研究结果总结如下：

我们发现笠贝Hox基因的表达的确呈现快速变化的特点，初步验证了我们的推测。这些变化与早期发育事件相关，包括原肠作用、贝壳发生、神经系统的发育及足的发生等。石鳖的Hox表达则随发育似乎变化不大。此外，与模式动物不同，笠贝和石鳖中Hox基因的背腹侧表达模式不一致。对多数Hox基因而言，它们可以只表达在腹侧或者背侧，又或者在两侧组织都有表达，但表达模式差异很大。基于这个特点，我们对背侧和腹侧的表达分别做了分析。在腹侧组织中，石鳖Hox基因的表达呈现出显著的共线性表达规律。在笠贝中，虽然其Hox基因的表达呈现快速变化，但是在早期担轮幼虫中，腹侧Hox的表达呈现出共线性模式。这是腹足纲软体动物Hox基因共线性表达的首次报道。这提示我们，软体动物都有Hox共线性表达模式，但是只局限在在特定的发育阶段的神经外胚层中。在背侧组织中，石鳖和笠贝的表达模式差异很大，但均与贝壳形成区相关。笠贝的Hox表达与幼虫圆形的贝壳相一致，多呈环形或圆形；石鳖Hox则多表达为重复的带状，与其重复排列的多个贝壳相符。

根据以上结果，我们提出一种普适的软体动物Hox基因表达模式。主要包括三点：（1）在腹侧神经外胚层存在保守的Hox基因共线性表达，（2）背侧Hox表达与贝壳形成区相关，体现出不同类群的独特性，（3）对有壳亚门动物，在发育晚期幼虫中，Hox基因主要表达在神经系统中。这种普适的Hox表达模型能够与大多数软体动物的Hox表达数据相适应，基本解决了软体动物Hox表达规律是否具有一致性以及是否存在共线性表达的争议。这些结果为理解软体动物的发育和演化提供了重要的基础支撑。例如，背腹分离的Hox基因表达模式提示软体动物的背腹侧发育也互相分离，从而各自在不同类群中发生多样化的演变，这可能是当前软体动物多样化体制的内在驱动机制。

Hox genes are important developmental regulatory genes and they play essential roles in body patterning of animals. Conserved Hox expression patterns are observed in a wide range of animals, which show staggered pattern along the anterior-posterior axis (spatial collinearity). However, such expression pattern is not observed in mollusks. Current research reveals that Hox expression shows very diverse patterns in different clades of mollusks, even among closely related species. Although limited evidence reveals staggered expression, it is observed in varied tissues among different mollusks. In addition, even without seeking a common staggered pattern, molluscan Hox expression is still too diverse to conclude a general model. Although correlations between Hox expression and larval shell field, foot and nervous system are frequently observed, the Hox expression in these particular organs shows few common patterns. Together, expression and functions of molluscan Hox genes are important issues to be addressed in the research field of development and evolution.

Based on these facts, we proposed that, there could have been dynamic Hox expression during molluscan development, which may lead to the diverse expression patterns observed in different developmental stages. To verify this speculation, in the present study, we focused on two phylogenetically distant mollusks, the gastropod Lottia goshimai (a limpet) and the polyplacophoran Acanthochitona rubrolineata (a chiton). We investigated the Hox expression in different developmental stages in the two molluscan species, aiming to find a generalized model of molluscan Hox expression, which may provide fundamental support for studies of molluscan development and evolution. The major results are described as follows.

We firstly confirmed that, as we had speculated, the Hox expression of L. goshimai indeed showed dynamic expression. The changes of Hox expression showed correlation with developmental events including gastrulation, shell formation, neurogenesis and foot development. Hox expression in A. rubrolineata showed minor changes at the two developmental stages examined. Moreover, unlike model organisms such as some insects and vertebrates, Hox expression in both L. goshimai and A. rubrolineata showed dissociated dorsal and ventral expression. At a given time point, a Hox gene could be solely expressed in dorsal or ventral tissues, or both, while showing distinct expression patterns. Therefore, we separated dorsal and ventral expression in subsequent analyses. On the ventral side, Hox expression of A. rubrolineata showed clear staggered pattern. In L. goshimai, despite the dynamic changes, we found a developmental stage, i.e., the early trochophore stage (10 hpf), when the ventral Hox expression showed staggered pattern. This is the first report showing staggered Hox expression in a gastropod mollusk. It indicates that molluscan Hox expression does show staggered pattern, which however is restricted to neuroectoderm at a particular developmental stage. On the dorsal side, although distinct Hox expression was observed in the two species, it showed high correlation with shell formation in a lineage-specific manner. In accordance with a round larval shell in L. goshimai, the dorsal Hox expression often showed a circle pattern surrounding the shell field or was restricted to the central region of the shell field. In A. rubrolineata, dorsal Hox expression was often observed as repeated bands, which coincided with its metameric larval shell plates.

From our results, we deduce a generalized model of molluscan Hox expression based on dissociated dorsal and ventral expression. First, ventral Hox expression shows a conserved staggered pattern at particular developmental stages (i.e., during early neuroectoderm patterning). Second, dorsal Hox expression correlates with the shell field and is highly lineage-specific. Lastly, in later larval stages of conchiferans, most Hox genes are exclusively expressed in the nervous system (yet no longer in a staggered manner). By closely examining previous reports, we conclude that this generalized model is compatible with known Hox expression data from various molluscan clades. These results clarify the long-standing debates concerning whether mollusks share common Hox expression and whether they have staggered pattern. The results also provide fundamental supports for understanding molluscan development and evolution. For instance, the dorsoventral dissociation of Hox expression indicate a dorsoventrally dissociated dorsal and ventral patterning, which therefore permits lineage-specific dorsal and ventral patterning in different molluscan clades and may underpin the diversification of molluscan body plans.