Quéré I, Paul V, Rouillac C, Janbon C, London J, Demaille J, Kamoun P, Dufier J L, Abitbol M, Chassé J F
Faculté de Médecine Necker, Hôpital Necker Enfants Malades, CNRS URA 1335, 156 rue de Vaugirard, Paris, 75015, France.
Biochem Biophys Res Commun. 1999 Jan 8;254(1):127-37. doi: 10.1006/bbrc.1998.9079.
We report the cystathionine-beta synthase (CBS) gene expression pattern during early human embryogenesis (3 to 6 weeks post conception) by in situ hybridization and in fetal and adult tissue by Northern Blot analysis. Probes were chosen to recognize either the common sequence to all known CBS mRNAs or the sequences of two different major exons 1 issued of we have previously identified. We demonstrate by in situ hybridization that CBS is continuously expressed from the earliest stages studied (22 days post conception) during embryogenesis in the tissues of developing embryos which will after birth present clinical abnormalities in homocystinuria patients. It is expressed at an especially high level in the neural and cardiac systems until the liver primordium appears. In embryonic central nervous system, the whole neural tube and primary brain vesicles are labeled. Secondary brain vesicles labeling are dependent on the neuroepithelium differentiation. The ventricular layer of the rhombencephalon, cranial nerve nuclei and then after cerebellar cortex derived from rhombencephalon ventricular layer are strongly labeled. Thalamus and other derivatives of the diencephalon plate, the neuroblastic layer of the retina, lens and dorsal root ganglia are labeled. After 35 days post conception, CBS mRNAs was detected in endocardial cells and in cells derived from the neural crest of the heart and in particular developing mesodermic regions such as the primitive hepatocytes of the liver, mesonephros vesicles, various endocrine glands and developing bones. We could not detect tissue specificity of different probes at this embryonic stage. Northern blot analysis consistently detected mRNA species in fetal 25 weeks post conception brain, liver and kidney. The common cDNA probe revealed the 2.5 and 3.7 kb mRNA species from brain, liver and kidney. The exon 1b probe detected only the 2.5 kb mRNA and the exon 1c probe the 3.7 kb mRNA in these three tissues. In adult tissue, the 1b probe detected only the 2.5 kb mRNA and the 1c probe only the 3.7 kb mRNA in the liver.
我们通过原位杂交技术报告了人胚胎发育早期（受孕后3至6周）胱硫醚-β-合酶（CBS）基因的表达模式，并通过Northern印迹分析报告了其在胎儿及成人组织中的表达模式。我们选择了能识别所有已知CBS mRNA共同序列的探针，以及我们之前鉴定出的两个不同主要外显子1的序列。通过原位杂交我们证明，在胚胎发育过程中，从研究的最早阶段（受孕后22天）开始，CBS就在发育中胚胎的组织中持续表达，这些组织在出生后会在同型胱氨酸尿症患者中出现临床异常。在肝脏原基出现之前，它在神经和心脏系统中表达水平特别高。在胚胎中枢神经系统中，整个神经管和初级脑泡都被标记。次级脑泡的标记取决于神经上皮的分化。菱脑的室层、脑神经核，以及随后源自菱脑室层的小脑皮质被强烈标记。丘脑和间脑板的其他衍生物、视网膜的神经母细胞层、晶状体和背根神经节也被标记。受孕后35天，在内皮细胞、源自心脏神经嵴的细胞以及特别是发育中的中胚层区域，如肝脏的原始肝细胞、中肾小泡、各种内分泌腺和发育中的骨骼中检测到了CBS mRNA。在这个胚胎阶段，我们无法检测到不同探针的组织特异性。Northern印迹分析在受孕后25周的胎儿脑、肝和肾中持续检测到mRNA种类。通用cDNA探针在脑、肝和肾中检测到2.5 kb和3.7 kb的mRNA种类。外显子1b探针在这三种组织中仅检测到2.5 kb的mRNA，外显子1c探针仅检测到3.7 kb的mRNA。在成人组织中，1b探针在肝脏中仅检测到2.5 kb的mRNA，1c探针仅检测到3.7 kb的mRNA。
