IDENTIFICATION AND DEVELOPMENTAL EXPRESSION OF A NOVEL LOW-MOLECULAR-WEIGHT NEURONAL INTERMEDIATE FILAMENT PROTEIN EXPRESSED IN XENOPUS-LAEVIS
IDENTIFICATION AND DEVELOPMENTAL EXPRESSION OF A NOVEL LOW-MOLECULAR-WEIGHT NEURONAL INTERMEDIATE FILAMENT PROTEIN EXPRESSED IN XENOPUS-LAEVIS
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DOI：
10.1523/jneurosci.12-08-03010.1992
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发表时间：
1992-08-01
影响因子：
5.3
通讯作者：
GAINER, H
中科院分区：
文献类型：
--
作者：
CHARNAS, LR;SZARO, BG;GAINER, H
非洲爪蟾是研究脊椎动物神经胚胎发生的一个有价值的模型系统。然而，很少有表征良好的神经系统特异性分子标记可用于该生物的研究。利用小鼠低分子量神经丝蛋白（NF-L） cDNA探针，筛选了一个成年鼠脑cDNA文库，以鉴定神经元特异性的中间丝蛋白。对两种不同的神经元特异性中间丝蛋白克隆进行了分离和测序。其中一种编码爪蟾NF-L (XNF-L)，另一种编码一种新的神经元特异性爪蟾中间丝蛋白（XNIF），这种蛋白在发育过程中比XNF-L更早、更丰富地存在。XNIF包含一个中心棒结构域，具有IF蛋白特有的多个序列特征。XNF-L与小鼠NF-L非常相似，在杆状结构域具有77%的序列同一性，在尾结构域存在聚谷氨酸区域，这是IV型神经丝蛋白的特征。相比之下，XNIF在杆状结构域与小鼠NF-L的同源性仅为60%，在尾部结构域缺乏谷氨酸丰富的序列。与NF-L和其他神经丝蛋白相比，XNIF在头部和尾部结构域的序列一致性也非常低（约38%）（与α -连接蛋白头部结构域的一致性为45%）。在成年青蛙中，XNIF mRNA仅在神经系统中被Northern印迹检测到，而在神经元中，特别是在髓网状系统和脊髓中，通过原位杂交组织化学检测到。针对XNIF独特尾部区域的抗血清在神经系统细胞骨架制剂的Western blots中检测到一个独特的60 kDa条带，该XNIF免疫反应性集中在PNS轴突和背根神经节的小核周。相反，NF-L的免疫反应主要发生在背根神经节的大核周。在发育过程中，XNIF mRNA比XNF-L mRNA更丰富。XNIF mRNA在24期（26小时）首次被检测到，而XNIF - l在35/36期（50小时）稳定表达。在骨水泥腺、发育中的神经系统内的许多神经元细胞体和轴突束以及发育中的视网膜的所有细胞层中均可检测到XNIF的免疫反应性。这两种具有不同时间和空间表达模式的神经元特异性中间丝蛋白的可用性，应该为发育中的松鸡神经系统功能扰动提供新的标记和靶点。
Xenopus laevis is a valuable model system for the study of vertebrate neuroembryogenesis. However, very few well-characterized nervous system-specific molecular markers are available for studies in this organism. We screened a X. laevis adult brain cDNA library using a cDNA probe for mouse low molecular weight neurofilament protein (NF-L) in order to identify neuron-specific intermediate filament proteins. Clones for two distinct neuron-specific intermediate filament proteins were isolated and sequenced. One of these encoded for a Xenopus NF-L (XNF-L) and the other for a novel neuron-specific Xenopus intermediate filament protein (XNIF) that was present earlier and more abundantly than XNF-L during development.XNIF contained a central rod domain with multiple sequence features characteristic of IF proteins. The XNF-L was very similar to mouse NF-L, with a 77% sequence identity in the rod domain and the presence of a polyglutamic acid region in the tail domain, characteristic of type IV neurofilament proteins. In contrast, XNIF showed only 60% identity to mouse NF-L in the rod domain and lacked the glutamic acid-rich sequence in the tail domain. XNIF also had a very low (approximately 38%) sequence identity in the head and tail domains as compared to NF-L and other neurofilament proteins (45% identity to the head domain of alpha-internexin).In the adult frog, XNIF mRNA is detected by Northern blots only within the nervous system and by in situ hybridization histochemistry exclusively in neurons, particularly in the medullary reticular system and spinal cord. Antisera raised against the unique tail region of XNIF detected a single distinct 60 kDa band in Western blots of nervous system cytoskeletal preparations, and this XNIF immunoreactivity was concentrated in axons in the PNS and in small perikarya in the dorsal root ganglion. in contrast, NF-L immunoreactivity was principally in the large perikarya in the dorsal root ganglion. In development, XNIF mRNA appears more abundant than XNF-L mRNA in all premetamorphic stages examined. XNIF mRNA is first detectable at stage 24 (26 hr), whereas stable expression of XNF-L is at stage 35/36 (50 hr). XNIF immunoreactivity is detectable within the cement gland, within many neuronal cell bodies and axon tracts within the developing nervous system, and within all cellular layers of the developing retina. The availability of these two distinct neuron-specific intermediate filament proteins, with different temporal and spatial expression patterns, should provide new markers as well as targets for functional perturbation in the developing X. laevis nervous system.