Skinner Debra J, Brown Ryan H, Kuzoff Robert K, Gasser Charles S
Department of Molecular and Cellular Biology, University of California, Davis, Davis, CA, 95616, USA.
Present address: US Patent and Trademark Office, 400 Dulany St, Alexandria, VA 22314, USA.
BMC Plant Biol. 2016 Jun 27;16(1):143. doi: 10.1186/s12870-016-0835-z.
The INNER NO OUTER (INO) gene is expressed in the outermost cell layer of the outer integument of bitegmic ovules and is essential for this organ's growth. The role and cross-species functional conservation of INO orthologs were examined in members of the Solanaceae, which have unitegmic ovules. Unitegmy has evolved several times in disparate angiosperm lineages. INO expression has been observed in the outermost cell layers of all examined unitegmic ovules, but the functional role of INO in unitegmic ovules has not previously been evaluated.
INO orthologs were unambiguously identified in tobacco and tomato by sequence homology. Expression of the tomato INO gene was limited to the outer cell layer of the single integument indicating that this single integument has properties of the outer integument. Expression occurred only after integument initiation, later than observed in ovules of other examined angiosperms. Virus-induced knock-down of expression of the INO ortholog in tobacco inhibited growth of the outer cell layer of the integument leading to a decrease in both integument extension and curvature of the ovule. The altered ovules closely resemble those of the aberrant testa shape (ats) ino mutant combination in Arabidopsis where we see the effect of the ino mutation on a single fused integument produced by the ats mutation. Despite significant sequence identity and similar expression patterns, the tomato INO coding region was not able to complement the Arabidopsis ino mutant.
The similarity of effects of ino mutations on the unitegmic ovules of tobacco and the fused integuments of the Arabidopsis ats mutant show that: 1) INO orthologs play the same role in promoting integument growth in ovules of tobacco and Arabidopsis; and 2) the unitegmic ovules of tobacco (and hence other solanaceous species) are most likely the result of a congenital fusion of two ancestral integuments. Our results further indicate that INO has a conserved role in growth of the outermost cell layer of integuments. The curvature of solanaceous ovules is driven by unequal growth of the outer layers of the single integument that likely correspond to an ancestral outer integument.
“内无外”（INO）基因在咬生胚珠外珠被的最外层细胞层中表达，对该器官的生长至关重要。在具有合生胚珠的茄科植物成员中，研究了INO直系同源基因的作用和跨物种功能保守性。合生现象在不同的被子植物谱系中已经进化了好几次。在所有检测的合生胚珠的最外层细胞层中都观察到了INO的表达，但INO在合生胚珠中的功能作用此前尚未得到评估。
通过序列同源性在烟草和番茄中明确鉴定出INO直系同源基因。番茄INO基因的表达仅限于单珠被的外层细胞，这表明该单珠被具有外珠被的特性。表达仅在珠被起始后发生，比在其他检测被子植物胚珠中观察到的时间晚。病毒诱导烟草中INO直系同源基因表达的敲低抑制了珠被外层细胞的生长，导致珠被延伸和胚珠曲率均降低。改变后的胚珠与拟南芥异常种皮形状（ats）ino突变体组合的胚珠非常相似，在拟南芥中我们看到ino突变对由ats突变产生的单个融合珠被的影响。尽管存在显著的序列同一性和相似的表达模式，但番茄INO编码区不能互补拟南芥ino突变体。
ino突变对烟草合生胚珠和拟南芥ats突变体融合珠被的影响相似表明：1）INO直系同源基因在促进烟草和拟南芥胚珠珠被生长中起相同作用；2）烟草（以及其他茄科物种）的合生胚珠很可能是两个祖先珠被先天性融合的结果。我们的结果进一步表明，INO在珠被最外层细胞层的生长中具有保守作用。茄科胚珠的曲率是由单珠被外层的不均衡生长驱动的，这可能对应于祖先的外珠被。
