Differential regulation of alpha 2u globulin gene expression in liver, lachrymal gland, and salivary gland

The alpha 2u globulins, products of a highly homologous multigene family, are synthesized in the liver and submaxillary salivary glands of the rat. Although their precise function has not been ascertained, they are of interest because of the complex developmental and hormonal regulation of their tissue levels. We now report that alpha 2u globulin is synthesized in a third tissue of the rat, the extraorbital lachrymal gland. Immunocytochemical studies indicate that the distribution of alpha 2u globulin is more homogeneous in the lachrymal gland than in the liver or submaxillary gland. In situ hybridization to alpha 2u globulin RNA reveals specific signal only over the acinar cells of the lachrymal gland. Several different isoelectric forms of alpha 2u globulin are encoded by lachrymal gland mRNA. The major lachrymal and salivary gland isoforms are indistinguishable from one another, but more acidic than the hepatic isoforms. In addition, analysis of double-stranded cDNAs with a diagnostic restriction-enzyme pair detects no differences between the alpha 2u globulin mRNAs of lachrymal and salivary gland, but clearly distinguishes these from their hepatic counterparts. In spite of the similarity between the lachrymal and salivary gland alpha 2u globulin gene products, we find that the hormonal and developmental regulation of alpha 2u globulin expression differs markedly in these two tissues. In the liver, where a different subset of alpha 2u globulin genes is expressed, a third regulatory phenotype is observed.     

Tissue-specific control of alpha 2u globulin gene expression: constitutive synthesis in the submaxillary gland

Synthesis of alpha 2u globulin, previously thought to occur only in the male rat liver, has now been demonstrated in the submaxillary salivary gland. Unlike liver, submaxillary synthesis of alpha 2u globulin mRNA is constitutive--that is, independent of the endocrine state, age and sex. Liver and submaxillary alpha 2u globulin mRNAs are of similar size, and their 5' ends map to the same region of the gene. Isoelectric focusing of in vitro translation products revealed that submaxillary mRNA encodes a more acidic subset of alpha 2u globulins than does liver. Salivary alpha 2u globulin mRNA manifests 5% nucleotide divergence, encoding 20 amino acid substitutions, which specifies a more acidic polypeptide than its hepatic counterpart. Thus the liver and submaxillary gland synthesize alpha 2u globulin from different sets of genes that are subject to very different developmental and hormonal control.     

Identification of three protein disulfide isomerase members from Haemaphysalis longicornis tick

Three genes encoding putative protein disulfide isomerase (PDI) were isolated from the Haemaphysalis longicornis EST database and designed as HlPDI-1, HlPDI-2, and HlPDI-3. All three PDI genes contain two typical PDI active sites CXXC and encode putative 435, 499, and 488 amino acids, respectively. The recombinant proteins expressed in Escherichia coli all show PDI activities, and the activities were inhibited by a PDI-specific inhibitor, zinc bacitracin. Western blot analysis and real-time PCR revealed that three HlPDIs were present in all the developmental stages of the tick as well as in the midgut, salivary glands, ovary, hemolymph, and fatbody of adult female ticks, but the three genes were expressed at the highest level in the egg stage. HlPDI-1 is expressed primarily in the ovary and secondarily in the salivary glands. HlPDI-2 and HlPDI-3 are expressed primarily in the salivary gland, suggesting that the PDI genes are important for tick biology, especially for egg development, and that they play distinct roles in different tissues. Blood feeding induced significantly increased expression of HlPDI-1 and HlPDI-3 in both partially fed nymphs and adults. Babesia gibsoni-infected larval ticks expressed HlPDI-1 and HlPDI-3 2.0 and 4.0 times higher than uninfected normal larval ticks, respectively. The results indicate that HlPDI-1 and HlPDI-3 might be involved in tick blood feeding and Babesia parasite infection in ticks.     

Expression of three forms of thyroid hormone receptor in human tissues

At least two thyroid hormone receptor (hTR) genes are present in humans, but the significance of this multiplicity is unknown. These receptors could have differences in tissue distribution or possess different functions. We studied the distribution and abundance of three hTR mRNAs (hTR beta, hTR alpha 1, and hTR alpha 2) by Northern blot analysis. Three mRNAs were expressed in all tissues examined. hTR beta was strongly expressed in brain and prostate predominantly as a 10.0-kilobase (kb) mRNA. This mRNA was also expressed in thyroid and was much less abundant in liver, kidney, placenta, tonsil, and spleen. hTR alpha 1 is represented by two mRNAs with sizes of 6.0 and 3.2 kb. The 6.0-kb mRNA was constantly less abundant than the 3.2-kb mRNA. hTR alpha 2 was detected as a single mRNA with a size of 3.2 kb, using a probe unique for this mRNA. Both hTR alpha 1 and hTR alpha 2 were strongly expressed in brain, prostate, and thyroid and much less in other tissues. The relative amounts of the three hTR mRNAs were roughly parallel in each tissue. It is of interest that none of these hTRs was abundant in liver, which is the major thyroid hormone-responsive organ. Another hTR may be present in liver.     

Global analysis of gene expression profiles in the submandibular salivary gland of klotho knockout mice

Salivary dysfunction commonly occurs in many older adults and is considered a physiological phenomenon. However, the genetic changes in salivary glands during aging have not been characterized. The present study analyzed the gene expression profile in salivary glands from accelerated aging klotho deficient mice (klotho?/?, 4 weeks old). Microarray analysis showed that 195 genes were differentially expressed (z‐score > 2 in two independent arrays) in klotho null mice compared to wild‐type mice. Importantly, alpha2‐Na⁺/K⁺‐ATPase (Atp1a2), Ca²⁺‐ATPase (Atp2a1), epidermal growth factor (EGF), and nerve growth factor (NGF), which have been suggested to be regulators of submandibular salivary gland function, were significantly decreased. When a network was constructed from the differentially expressed genes, proliferator‐activated receptor‐γ (PPAR γ), which regulates energy homeostasis and insulin sensitivity, was located at the core of the network. In addition, the expression of genes proposed to regulate various PPAR γ‐related cellular pathways, such as Klk1b26, Egfbp2, Cox8b, Gpx3, Fabp3, EGF, and NGFβ, was altered in the submandibular salivary glands of klotho?/? mice. Our results may provide clues for the identification of novel genes involved in salivary gland dysfunction. Further characterization of these differentially expressed genes will be useful in elucidating the genetic basis of aging‐related changes in the submandibular salivary gland.

     

棉铃虫幼虫唾液腺cDNA文库的构建及EST分析

棉铃虫Helicoverpa armigera (Hübner)幼虫唾液中的各种酶类及各种生化组分在棉铃虫与植物相互作用及协同进化中起到重要作用; 唾液腺是棉铃虫唾液成分的合成器官。本研究通过构建棉铃虫幼虫唾液腺全长cDNA文库, 测序得到1 502条EST序列, 聚类分析后获得821个unigenes, 为筛选棉铃虫与寄主互作信号因子提供基因信息资源。使用Blast2 GO软件对821个unigenes进行了比对和功能注释, 初步获得棉铃虫幼虫唾液腺中mRNA的构成特征。结果显示, 在棉铃虫唾液腺ESTs文库中, 鉴定得到脂类相关消化酶基因17个, 糖类相关消化酶基因5个, 半胱氨酸蛋白酶基因1个, 丝氨酸蛋白酶基因20个（其中16个为新发现）, 提示唾液腺的主要功能是分泌消化酶进行预消化; 还发现在棉铃虫幼虫唾液腺中存在表皮蛋白、 气味结合蛋白和化学感受蛋白基因。结果为研究棉铃虫预消化系统打下基础。     

Expression of procyclin mRNAs during cyclical transmission of Trypanosoma brucei

Trypanosoma brucei, the parasite causing human sleeping sickness, relies on the tsetse fly for its transmission. In the insect, EP and GPEET procyclins are the major surface glycoproteins of procyclic (midgut) forms of the parasite, with GPEET predominating in the early procyclic form and two isoforms of EP in the late procyclic form. EP procyclins were previously detected on salivary gland trypanosomes, presumably epimastigotes, by immunoelectron microscopy. However, no procyclins could be detected by mass spectrometry when parasites were isolated from infected glands. We have used qualitative and quantitative RT-PCR to analyse the procyclin mRNAs expressed by trypanosomes in the tsetse midgut and salivary glands at different time points after infection. The coding regions of the three EP isoforms (EP1, EP2 and EP3) are extremely similar, but their 3' untranslated regions contain unique sequences that make it possible to assign the cDNAs amplified by this technique. With the exception of EP2, we found that the spectrum of procyclin mRNAs expressed in the midgut mirrors the protein repertoire of early and established procyclic forms. Surprisingly, procyclin mRNAs, including that of GPEET, are present at relatively high levels in salivary gland trypanosomes, although the proteins are rarely detected by immunofluorescence. Additional experiments using transgenic trypanosomes expressing reporter genes or mutant forms of procyclin point to a mechanism of translational or post-translational control, involving the procyclin coding regions, in salivary gland trypanosomes. It is widely accepted that T. brucei always has a coat of either variant surface glycoprotein or procyclin. It has been known for many years that the epimastigote form does not have a variant surface glycoprotein coat. The finding that this life cycle stage is usually negative for procyclin as well is new, and means that the paradigm will need to be revised.     

Early events in the steroidal regulation of alpha2mu globulin in rat liver. Evidence for both androgenic and estrogenic induction.

1. A double-antibody radioimmunoassay for alpha2mu globulin has been developed. With the help of this highly sensitive radoimmunoassay the early effects of both androgen and estrogen treatments on the hepatic synthesis of alpha2mu globulin in the rat have been investigated. 2. Results show that the earlier observation of the long lag period in the androgenic induction of alpha2mu globulin is more apparent than real. 3. Single injections of either 5alpha-dihydrotestosterone(5alpha-dihydro-17beta-hydroxy-4-androsten-3-one) or its physiological antagonist estradiol-17beta (1,3,5(10)-estratriene-3,17beta-idol) to castrated female rats resulted in the induction of alphamu globulin reaching maximum hepatic level of the protein between 6--9 h after the hormone administration. Administration of cycloheximide 15 prior to hormone treatment blocked both androgenic andestrogenic induction of alpha2mu globulin. 4. Daily pretreatments with 5alphamu-dihydrotestosterone increased the sensitivity of subsequent androgenic response to alpha2muglobulin synthesis. On the other hand, daily pretreatments with estradiol 17beta decreased and ultimately abolished the estrogenic induction of alpha2mu globulin. 5. The possible mechanism of both androgenic and estrogenic induction of alpha2mu globulin in rat liver mediated through a sex-hormone-binding protein with dual affinity for both dihydrotestosterone and estradiol has been suggested.