Global chimeric exchanges within the intracellular face of the bradykinin B2 receptor with corresponding angiotensin II type Ia receptor regions: generation of fully functional hybrids showing characteristic signaling of the AT1a receptor

The intracellular (IC) face of the G-protein coupled receptors (GPCR), bradykinin (BK) B2 and angiotensin (AT) 1a, is similar in sequence homology and in size. Both receptors are known to link to Galphai and Galphaq but differ markedly in a number of physiologic actions, particularly with respect to their hemodynamic action. We made single as well as multiple, global replacements within the IC of BKB2R with the corresponding regions of the AT1aR. When stably transfected into Rat-1 cells, these hybrid receptors all bound BK with high affinity. Single replacement of the intracellular loop 2 (IC2) or the distal 34 residues of the C-terminus (dCt) with the corresponding regions of AT1aR resulted in chimera, which turned over phosphotidylinositol (PI) and released arachidonic acid (ARA) as WT BKB2R. In contrast, incorporation of the AT1aR IC3 in a single replacement abolished signal transduction. However, the simultaneous exchange of IC2 and IC3 of BKB2R with AT1aR resulted in a receptor responding to BK with PI turnover and ARA release approximately 4-fold greater than WT BKB2R. Likewise, the simultaneous replacement of IC2 and dCt resulted in a 2.8- and 1.6-fold increase in PI turnover and ARA release, respectively. In contrast, the dual replacement of IC3 and dCt could not overcome the deleterious effects of the IC3 replacement, resulting in very low PI activation and ARA release. Replacement of all three IC domains (IC2, IC3, and dCt) resulted in PI closer to that of AT1aR than BKB2R. The uptake of the receptor chimeras was similar to that of WT BKB2R with the exception of the IC3/dCt dual mutant, which exhibited very poor internalization (18% at 60'). When transfected into Rat-1 cells, the AT1aR markedly increased the expression of connective tissue growth factor (CTGF) mRNA, while BK slightly decreased it. The dual IC2/dCt and triple IC2/IC3/dCt hybrids both upregulated CTGF mRNA in response to BK. These results show that the IC face of the BKB2R can be exchanged with that of AT1aR, producing hybrid receptors, which take on the functional characteristics of AT1aR. The characterization of the chimera with stepwise replacement of the IC domains should allow for assignment of specific roles to the individual loops and C-terminus in the signaling and internalization of the BKB2R and facilitate the generation of a receptor with BKB2R binding and AT1aR function.     

人恶性疟杂合多肽抗原基因化学合成及克隆

本文报道用固相亚磷酰胺法合成人恶性疟杂合多肽抗原基因。基因全长为216bp,分为10个寡聚核苷酸片段分别合成,然后经T4 DNA连接酶按设计顺序连接成完整的杂合抗原基因,重组到噬茵体M13 mp 18 RF DNA内,转染大肠杆菌JM109。用分子杂交和酶切分析筛选出重组克隆体。经序列分析,证明所合成的人恶性疟杂合多肽抗原基因与所设计完全一致。     

In situ hybridization with species-specific DNA probes gives evidence for asymmetric nature of Brassica hybrids obtained by X-ray fusion

Summary We have previously reported production of somatic hybrids between B. oleracea and B. campestris by fusion of B. oleracea protoplasts with X-irradiated B. campestris protoplasts, in order to transfer a part of the B. campestris genome into B. Oleracea. Our previous analysis of morphology, chromosome number, and isozyme patterns of the hybrids suggested that they are asymmetric in nature. To obtain further evidence for the asymmetric nature of the hybrids, we isolated B. campestris-specific repetitive sequences and used them for in situ hybridization of the chromosomes of the hybrids. The repetitive DNA probes could specifically identify 8 out of 20 chromosomes of the B. campestris genome, and analysis of the hybrids indicates that 1–3 chromosomes of B. campestris are lacking in all five hybrids examined, giving clear evidence for the asymmetric nature of the hybrids. Furthermore, in situ hybridization revealed that some of the abnormal chromosomes observed in the hybrids are generated by rearrangements of B. Campestris chromosomes caused by X-irradiation. Altogether, our study indicates that in situ hybridization using species-specific repetitive sequences is a useful tool to analyze chromosomal compositions of various types of hybrids obtained by cell fusion or conventional methods.     

In vitro flower formation from thin epidermal cell layers of a partial somatic hybrid between Petunia hybrida (Hort.) and Nicotiana plumbaginifolia (Viv.)

In vitro flowers have been obtained by culturing thin epidermal cell layers of a partial somatic intergeneric hybrid. The phenotype of these flowers differs from that of flowers formed on seed-grown plants (in situ flowers) and from that of flowers of either parental line. In addition, modifications in the phenotype were observed when cultures were sustained for more than four months. Dimorphic leaves present in juvenile and adult stages of mother plants of Nicotiana plumbaginifolia and the somatic hybrid were formed on different ends of the thin epidermal cell layers. No anomalies were observed during microsporogenesis and in the meiotic and mitotic figures of the somatic hybrid, which resembled those of Nicotiana plumbaginifolia.     

东北马鹿和东北梅花鹿F_1杂种精母细胞联会复合体分析

作者以界面铺张-硝酸银染色技术,对东北马鹿和东北梅花鹿的F_1可育杂种的精母细胞联会复合体进行亚显微观察及分析。在减数分裂前期,杂种鹿精母细胞中形成31条完整的常染色体联会复合体、一个端着丝粒染色体/中着丝粒染色体的三价体和XY双价体。这进一步证明,两种亲本鹿的染色体具有高度的同源性,其差别仅在于一个罗伯逊易位。三价体的顺式构型可能和杂种鹿的可育性有关。     

水稻籼粳杂种雌性不育的细胞学初步观察(简报)

普通栽培稻籼粳亚种间杂种结实率低是开展亚种间杂交育种和杂种优势利用的主要障碍。这一障碍是杂种花粉和胚囊的不育性引起的。不育性曾经认为是两者染色体在结构上存在微小的差异所致,但F_1植株减数分     

Molecular evidence for mating asymmetry and female choice in a pocket gopher (Thomomys) hybrid zone

This paper presents circumstantial evidence that the mating system of the North American pocket gophers (Rodentia: Geomyidae) is a promiscuous one, with female choice at its base. A molecular marker (a length variant in the mitochondrial Control region [D-loop]) is used to show mating asymmetry in a hybrid zone between the species Thomomys bottae and Thomomys townsendii in north-eastern California. All hybrids result from a bottae mother × townsendii father cross. Because of significant differences in body size and resulting burrow diameter, bottae females must have actively sought their respective townsendii mates for the asymmetry in mating to have occurred, signalling female choice in these subterranean mammals that are otherwise characterized by exclusive-use territories, skewed adult sex ratio in favour of females, and high variance in male reproductive success.     

长穗偃麦草细胞质小麦核质杂种农艺性状的研究

长穗偃麦草细胞质小麦核质杂种与核共体进行了比较试验,结果证明：核质杂种株高降低５－８ｃｍ,抽穗期略有推迟,抗寒性有明显提高。其它农艺性状,如分蘖数,穗长,穗粒数,结实率等性状没有明显差异。但是在核质杂种中粒粒蛋白质含量增加了１１．７９％和２７．３３％,被分析的１７种氨基酸含量也是明显提高,提高幅度１２．３７－５３．０９％。     

Establishment of somatic hybrid cell lines between Zea mays L. (maize) and Triticum sect,trititrigia MacKey (trititrigia)

Summary Somatic hybrid cell lines were constructed by the fusion of protoplasts isolated from cell suspensions of Zea mays L. (maize, 2n = 20) and Triticum sect, trititrigia MacKey (trititrigia, 2n = 35), a perennial hybrid of T. durum Desf. and Elytrigia intermedium (Host) Nevski. Iodoacetamide-inactivated protoplasts of maize were fused with trititrigia protoplasts, which were sensitive to the PEG/DMSO fusion treatment at high pH and high calcium. Based on physiological complementation, approximately 0.002% of the total protoplasts cultured following fusion treatment developed into cell colonies, and 79 lines of them, almost a half, were singled out and subcultured. Among the subcultured lines three were, in comparison with the parents, identified as somatic hybrids by their coupled XbaI restriction patterns of total DNAs probed with the ribosomal DNA of rice. Southern analysis of the digested total DNAs with a mitochondrial gene, atpA., from pea, or a chloroplast gene, trnK, from rice, revealed that all the hybrids carried only the organellar DNAs of trititrigia, which excluded the possibilities of a chimeric callus or any DNA contamination. Cytogenetically, one hybrid was mixoploid with a 2n of 46–67 in which chromosomal endoreduplication, characterized by the appearance of diplochromosomes, was occasionally observed. Its hybridity was reconfirmed by the fact that it bore the satellite chromosomes of both maize and trititrigia, which were distinguishable from each other by size. In contrast, the other two hybrids were aneuploids. The potential of gene transfer between Zea and Triticum species was thus conclusively established.