Mínguez A, Moreno Díaz de la Espina S
Laboratorio de Biología Celular y Molecular Vegetal, Centro de Investigaciones Biológicas, Madrid, Spain.
J Cell Sci. 1993 Sep;106 ( Pt 1):431-9. doi: 10.1242/jcs.106.1.431.
We have used polyclonal and monoclonal antibodies against different lamins from vertebrates, and the IFA antibody recognizing all kinds of intermediate filament proteins, to investigate the lamins of the nuclear matrix of Allium cepa meristematic root cells. All the antibodies react in the onion nuclear matrix with bands in the range of 60-65 kDa, which are enriched in the nuclear matrix after urea extraction, and do not crossreact with other antibodies recognizing intermediate filaments in plants (AFB, anti-vimentin and MAC 322), ruling out crossreaction with contaminating intermediate filaments of cytoplasmic bundles. In 2-D blots the chicken anti-lamin serum reacts with one spot at 65 kDa and pI 6.8 and the anti B-type lamin antibodies with another one at 64 kDa and pI 5.75. Both crossreact with IFA. The lamin is localized at the nuclear periphery and the lamina by indirect immunofluorescence. Immunogold labelling of nuclear matrix sections reveals that the protein is not only associated with the lamina, but also with the internal matrix. Taken together these results reveal that higher plants, which do not possess an organized network of cytoplasmic intermediate filaments, nevertheless present a well-organized lamina containing lamins in which at least one of them is immunologically related to vertebrate lamin B. Our data confirm that lamins are very old members of the intermediate filament proteins that have been better conserved in plants during evolution than their cytoplasmic counterparts.
我们使用了针对脊椎动物不同核纤层蛋白的多克隆和单克隆抗体，以及识别各类中间丝蛋白的免疫荧光抗体（IFA），来研究洋葱根尖分生组织细胞的核基质中的核纤层蛋白。所有抗体在洋葱核基质中均与60 - 65 kDa范围内的条带发生反应，这些条带在尿素提取后在核基质中富集，并且不与识别植物中其他中间丝的抗体（AFB、抗波形蛋白和MAC 322）发生交叉反应，排除了与细胞质束中污染的中间丝发生交叉反应的可能性。在二维印迹中，鸡抗核纤层蛋白血清与一个位于65 kDa、pI 6.8的斑点发生反应，抗B型核纤层蛋白抗体与另一个位于64 kDa、pI 5.75的斑点发生反应。两者均与免疫荧光抗体（IFA）发生交叉反应。通过间接免疫荧光法，核纤层蛋白定位于核周边和核纤层。核基质切片的免疫金标记显示，该蛋白不仅与核纤层相关，还与内部基质相关。综合这些结果表明，高等植物虽然不具备有组织的细胞质中间丝网络，但仍存在一个组织良好的含有核纤层蛋白的核纤层，其中至少有一种核纤层蛋白在免疫上与脊椎动物的核纤层蛋白B相关。我们的数据证实，核纤层蛋白是中间丝蛋白中非常古老的成员，在植物进化过程中比其细胞质对应物得到了更好的保守。
