DesGroseillers L, Cowan D, Miles M, Sweet A, Scheller R H
Department of Biological Sciences, Stanford University, California 94305.
Mol Cell Biol. 1987 Aug;7(8):2762-71. doi: 10.1128/mcb.7.8.2762-2771.1987.
Neuropeptide genes are expressed in specific subsets of large polyploid neurons in Aplysia californica. We have defined the transcription initiation sites of three of these neuropeptide genes (the R14, L11, and ELH genes) and determined the nucleotide sequence of the promoter regions. The genes contain the usual eucaryotic promoter signals as well as other structures of potential regulatory importance, including inverted and direct repeats. The L11 and ELH genes, which are otherwise unrelated, have homology in the promoter regions, while the R14 promoter was distinct. When cloned plasmids were microinjected into Aplysia neurons in organ culture, transitions between supercoiled, relaxed circular, and linear DNAs occurred along with ligation into high-molecular-weight species. About 20% of the microinjected neurons expressed the genes. The promoter region of the R14 gene functioned in expression of the microinjected DNA in all cells studied. When both additional 5' and 3' sequences were included, the gene was specifically expressed only in R14, suggesting that the specificity of expression is generated by a multicomponent repression system. Finally, the R14 peptide could be expressed in L11, demonstrating that it is possible to alter the transmitter phenotype of these neurons by introduction of cloned genes.
神经肽基因在加州海兔的大型多倍体神经元的特定亚群中表达。我们已经确定了其中三个神经肽基因（R14、L11和ELH基因）的转录起始位点，并测定了启动子区域的核苷酸序列。这些基因包含常见的真核启动子信号以及其他具有潜在调控重要性的结构，包括反向和正向重复序列。L11和ELH基因在其他方面没有关联，但在启动子区域具有同源性，而R14启动子则不同。当将克隆质粒显微注射到器官培养的海兔神经元中时，超螺旋、松弛环状和线性DNA之间会发生转变，并连接成高分子量物种。大约20%的显微注射神经元表达了这些基因。R14基因的启动子区域在所研究的所有细胞中都能在显微注射DNA的表达中发挥作用。当同时包含额外的5'和3'序列时，该基因仅在R14中特异性表达，这表明表达的特异性是由多组分抑制系统产生的。最后，R14肽可以在L11中表达，这表明通过引入克隆基因有可能改变这些神经元的递质表型。
