王浆主蛋白MRJP7基因在意大利蜜蜂体内的表达

王浆蛋白是蜂王浆生物功能的物质基础,是由王浆蛋白基因家族(mrjps)编码合成的。但部分家族成员如MRJP7在王浆中的含量极少甚至检测不到。基因功能与其在生物体内的时空表达特性相关,为探究mrjp7的生物学功能,本研究利用荧光定量PCR技术对mrjp7在不同发育时期的工蜂和成年工蜂、雄蜂和蜂王的不同组织部位的表达进行定量检测。结果显示mrjp7在成年雄蜂体内的表达水平最低,成年蜂王次之,且在它们的各不同组织部位之间的表达量差异较小。该基因在工蜂幼虫和蛹期的表达同样较低,但在羽化后9日龄前后的哺育蜂王浆腺和头部特异性高表达,这与哺育蜂分泌蜂王浆哺育幼虫和蜂王的功能是相适应的,该结果在转录水平上证实了mrjp7的营养功能,为进一步的研究和应用打下了理论基础。     

蜜蜂蜂王浆主蛋白(MRJPs)的研究进展

蜜蜂的蜂王浆主蛋白具有为蜂王和幼虫提供营养、影响蜂群社会行为及调节个体生理机能等作用,作为蜂王浆的主要成分对其他机体也可产生多方面的生物学功能。因此,近年来蜂王浆主蛋白的相关研究备受关注。本文针对蜂王浆主蛋白的发现、种类、功能、系统进化及其基因表达情况进行了系统综述。     

蜜蜂体内王浆主蛋白基因mrjp9的转录表达研究

蜜蜂体内有9种王浆蛋白基因(major royal jelly protein,MRJPs1～9),其中MRJPs1～5在蜂王浆中含量较高,是蜂王浆生物学功能的基础。MRJPs6～9在王浆中没有或含量极少,且功能未知。为研究非王浆蛋白组分的MRJP9的生物学功能,本研究用RT-PCR的方法对意大利蜜蜂Apis mellifera ligustica Spinola不同组织,不同部位,不同级型样本中mrjp9的转录水平进行检测和定量。结果发现mrjp9在蜜蜂的幼虫、蛹和成年蜜蜂的各组织部位均广泛转录表达,但其在幼虫、蛹和刚出房的成年蜜蜂体内表达水平较低,而在成年采集蜂体内表达水平则较高,其表达与蜜蜂的发育时期有关。通过对在成年蜜蜂体内各组织部位的表达水平进行检测的结果显示该基因主要在蜜蜂的头、胸和王浆腺等组织部位的表达较高,其他组织部位表达较少。此外,该基因也在雄蜂和蜂王体内广泛表达,不受蜜蜂性别和级型的影响。这些结果说明mrjp9是一与蜜蜂发育有关的基因,可能与蜜蜂的行为发育和分工调控有关。     

蜂王浆主蛋白研究进展

作为社会性昆虫,蜜蜂是研究社会行为和学习记忆的理想模式生物。王浆主蛋白（Major royal jelly protein, MRJP）是蜂王浆中蛋白质的主要成分,该家族一共有9个成员,MRJP1~MRJP9。所有mrjps均以串联排列的形式位于蜜蜂11号染色体上一个大约60 kb的DNA片段上。mrjp的同源体也存在于其他的膜翅目昆虫,均是通过yellow进化而来的。随着不断地进化,MRJPs家族进化出许多重要功能,其中最主要的就是营养功能。本文从MRJPs家族的基因及蛋白质结构、mRNA表达情况、进化和功能等方面进行综述,为今后开展相关研究提供理论支持。     

Proteomic analysis of honeybee (Apis mellifera L.) pupae head development

The honeybee pupae development influences its future adult condition as well as honey and royal jelly productions. However, the molecular mechanism that regulates honeybee pupae head metamorphosis is still poorly understood. To further our understand of the associated molecular mechanism, we investigated the protein change of the honeybee pupae head at 5 time-points using 2-D electrophoresis, mass spectrometry, bioinformatics, quantitative real-time polymerase chain reaction and Western blot analysis. Accordingly, 58 protein spots altered their expression across the 5 time points (13-20 days), of which 36 proteins involved in the head organogenesis were upregulated during early stages (13-17 days). However, 22 proteins involved in regulating the pupae head neuron and gland development were upregulated at later developmental stages (19-20 days). Also, the functional enrichment analysis further suggests that proteins related to carbohydrate metabolism and energy production, development, cytoskeleton and protein folding were highly involved in the generation of organs and development of honeybee pupal head. Furthermore, the constructed protein interaction network predicted 33 proteins acting as key nodes of honeybee pupae head growth of which 9 and 4 proteins were validated at gene and protein levels, respectively. In this study, we uncovered potential protein species involved in the formation of honeybee pupae head development along with their specific temporal requirements. This first proteomic result allows deeper understanding of the proteome profile changes during honeybee pupae head development and provides important potential candidate proteins for future reverse genetic research on honeybee pupae head development to improve the performance of related organs.     

感染囊幼病的工蜂咽下腺细胞的超微结构的变化

前言 中华蜜蜂囊幼病是我国养蜂业的一种重要病毒病,发病率很高,有的地区造成大量的幼虫死亡,给我国养蜂生产带来一定的危害,为了对防治病害提供科学的依据,我们首先对该病病原进行了分离、提纯及电子显微镜的研究等工作。发现该病毒对幼虫和成蜂,尤其对工蜂体内各个器官都有程度不同的影响。在幼虫发病期症状特别明显,病幼虫身体松软多水,其表皮容易破裂,当悬挂幼虫时其幼虫末端积聚有透明的液滴,在巢房内幼虫头部尖并变成黑色,头部稍微向上抬起呈船形。死后的幼虫干涸变为褐色的外壳留于巢房内,当感染的幼虫全部封盖后,巢房盖的中央有一小孔,这些异常的变化都是囊幼病的典型症状。幼虫症状虽很明显,但工蜂感染病毒后在外部形态上却没有明显     

西方蜜蜂不同级型王浆主蛋白MRJP8基因的表达差异

王浆主蛋白在蜜蜂的级型分化中具有重要的功能。为探究mrjp8在西方蜜蜂Apis mellifera不同级型的表达模式及功能差异。【方法】 利用荧光定量PCR技术对西方蜜蜂工蜂、 雄蜂和蜂王不同发育时期和不同组织的mrjp8表达水平进行检测。【结果】 工蜂体内mrjp8在9日龄前后的毒腺组织内特异性高表达, 为参照基因表达量的上万倍, 在其他发育时期和组织的表达量则明显较低, 其表达具有明显的时空特异性; 在雄蜂体内其表达量与对照相当; 在蜂王体内表达量可达参照的近1 000倍, 没有组织特异性。【结论】 mrjp8的这种表达模式提示其在工蜂防御及维系蜂王长寿命方面有积极作用, 这为进一步研究该基因乃至整个王浆蛋白基因家族的进化和功能分化提供了依据。     

意蜂王浆生产性能形态学遗传标记的研究

测定19只工蜂头部咽下腺的两条腺体长度和重量,经生统t检验,发现同一工蜂的左、右两条咽下腺长度、重量差异都不显著（P＜0.05）。测定11群 "浙农大1号"意蜂(Apis mellifera ligustica)的330只工蜂的头重、体重、头体重比、咽下腺小囊数和咽下腺长度。在2001年3月30日至4月26日浙江淳安的油菜和紫云英花期测定试验蜂群的王浆产量,分析工蜂头重、头体重比、咽下腺小囊数和咽下腺长度与每千克蜂的王浆产量之间的相关性,发现咽下腺长度与王浆产量的相关性最大,小囊数次之,咽下腺长度可作为意蜂王浆生产性能较理想的形态学遗传标记。 Abstract:The lengths of hypopharyngeal glands (HG) from the left and right side were determined in 19 workers of honeybee(Apis mellifera ligustica).There were no significant differences (P＜0.05) in length between the left and the right in one worker′s hypopharyngeal gland.Three hundred and thirty workers were collected from eleven colonies of "ZND No.1" Italian honeybee(Apis mellifera ligustica) respectively.Head weight,body weight,ratio between head weight and body weight,bursa number and length of hypopharyngeal gland(HG)were tested in these samples.Royal jelly productions were determined during the flowing period of rape and Chinese milk retch from March 30 to April 26 in Chun′an County of Zhejiang Province in 2001.The correlation analysis between royal jelly production and head weight,ratio between head weight and body weight,bursa number of HG,and length of HG were conducted.The correlation coefficient between royal jelly production and length of HG was the largest.The correlation coefficient between royal jelly production and bursa number was the second.It was suggested that the length of HG could be used as one of genetic markers for the production performance of royal jelly.     

Identification of major royal jelly proteins in the brain of the honeybee Apis mellifera

The consumption of royal jelly (RJ) determines the differences between castes and behavioral development in the honeybee Apis mellifera. However, it is not known whether the proteins of RJ are related to these differences, or which proteins are responsible for the changes. To understand the functions of RJ proteins that are present in other tissues of the bee, in addition to hypopharyngeal gland, we used a polyclonal antibody anti-MRJP1 to investigate the presence of this protein in nervous system of honeybee. This study showed the presence of three polypeptides (p57, p70 and p128) in specific tissues of bee brain. Mushroom body, optic lobe and antennal lobe neuropils all contained proteins recognized by anti-MRJP1. Proteomic analysis showed that the three polypeptides are correlated with proteins of the MRJP family. p57 is correlated with MRJP1, p70 with MRJP3, while p128 may be an oligomeric form or a new polypeptide. Immunostaining of the brain and hypopharyngeal gland revealed differential expression of MRJPs in various brain regions and in different honeybee castes and subcastes. The identification and localization of these MRJPs contribute to the elucidation of the biological roles of this protein family.     

Origin and function of the major royal jelly proteins of the honeybee (Apis mellifera) as members of the yellow gene family

In the honeybee, Apis mellifera, the queen larvae are fed with a diet exclusively composed of royal jelly (RJ), a secretion of the hypopharyngeal gland of young worker bees that nurse the brood. Up to 15% of RJ is composed of proteins, the nine most abundant of which have been termed major royal jelly proteins (MRJPs). Although it is widely accepted that RJ somehow determines the fate of a female larva and in spite of considerable research efforts, there are surprisingly few studies that address the biochemical characterisation and functions of these MRJPs. Here we review the research on MRJPs not only in honeybees but in hymenopteran insects in general and provide metadata analyses on genome organisation of mrjp genes, corroborating previous reports that MRJPs have important functions for insect development and not just a nutritional value for developing honeybee larvae.     

Oral Administration of Royal Jelly Restores Tear Secretion Capacity in Rat Blink-Suppressed Dry Eye Model by Modulating Lacrimal Gland Function

Tears are secreted from the lacrimal gland (LG), a dysfunction in which induces dry eye, resulting in ocular discomfort and visual impairment. Honey bee products are used as a nutritional source in daily life and medicine; however, little is known about their effects on dry eye. The aim of the present study was to investigate the effects of honey bee products on tear secretion capacity in dry eye. We selected raw honey, propolis, royal jelly (RJ), pollen, or larva from commercially available honey bee products. Tear secretion capacity was evaluated following the oral administration of each honey bee product in a rat blink-suppressed dry eye model. Changes in tear secretion, LG ATP content, and LG mitochondrial levels were measured. RJ restored the tear secretion capacity and decrease in LG ATP content and mitochondrial levels to the largest extent. Royal jelly can be used as a preventative intervention for dry eye by managing tear secretion capacity in the LG.     

Conservation of nutrients in larval tissue by cannibalizing honey bees

ABSTRACT. ¹⁴C-phenylalanine appeared in royal jelly secreted by worker honey bees ( Apis mellifera , L.) directly after eating ¹⁴-phenyl-alanine labelled larvae. The amount of labelled phenylalanine in royal jelly was associated directly with the volume of jelly produced. Loss of radio-label from the workers, in the form of O₂¹⁴C, increased when royal jelly secretion ended. These patterns show that conservation of nutrients in cannibalized tissue is enhanced when the cannibal bees are producing royal jelly. Specific activity of radiolabel in worker bee hypopharyngeal glands was higher than in thoracic muscle, showing selective movement within the bees of a nutrient acquired by cannibalism.     

The MRJP/YELLOW protein family of Apis mellifera: identification of new members in the EST library

Major royal jelly proteins (named MRJP1-5) of honeybee (Apis mellifera), yellow proteins of Drosophila, together with putative proteins found in several bacteria, form a protein family termed the MRJP/yellow family. Members of the family exert diverse physiological functions and amongst eukaryotes appear to be restricted to the order Insecta. MRJPs constitute about 90% of total protein of royal jelly, which is secreted by nurse bees to feed the queen and growing larvae. We looked for mrjp and yellow homologues in a honeybee brain expressed sequence tags (EST) library. In addition to the five mrjp cDNAs previously characterized, we found three additional cDNAs encoding novel MRJPs and importantly, two cDNAs coding for orthologues of Drosophila yellow proteins. One yellow cDNA and all three cDNAs coding for the novel MRJPs were assembled completely, the sequence of the other yellow homologue was partially assembled. The data we present here supports the view that repeated duplications and functional divergence occurred during the evolution of MRJPs in honeybees, with even closely related MRJPs appearing to perform diverse physiological functions. Conversely, yellow protein orthologues appear to be conserved and thus candidates for maintaining the former function(s) of yellow proteins.     

The influence of some honeybee products as a diet substitute on the different stages of Coccinella undecimpunctata L. in Egypt

Several natural diets of honeybee products were tested for rearing Coccinella undecimpunctata L. under laboratory conditions. Two groups of tested adult and larval insect have been fed on the tested diets, the first group were fed on the tested honey bee products diets without adding aphids, while the second group fed on the same honeybee products with aphids added. The oviposition period in the first group was significantly shorter than control. The tested diets supplied with royal jelly + aphid and/or pollen grains + aphid caused the longest oviposition period. Diets of aphid alone and bee honey with aphid recorded the shortest oviposition period. Fecundity of ladybird was the highest in the tested diets with aphids than diets without aphid and/or aphid diet alone. The durations of larval and pupal stages were significantly different when compared with the diets with or without aphid and/or aphid diet alone. Further results indicated that the diets of royal jelly caused the shortest duration in larval and pupal stages.     

Antimicrobial activity of the C-terminal of the major royal jelly protein 4 in a honeybee (Apis cerana)

The protein component of honeybee royal jelly (RJ) is constituted by major royal jelly proteins (MRJPs). The Asiatic honeybee (Apis cerana) MRJP-4 (AcMRJP4) exhibits antimicrobial activities. In this study, we identified the antimicrobial activity of AcMRJP4-15, which is a hydrophilic peptide with 88 amino acid residues in the C-terminal of AcMRJP4 that contains a high content of Asn and positively charged amino acids. Recombinant AcMRJP4-15, which is expressed as a 15-kDa peptide in baculovirus-infected insect cells, induced structural damage to the cell walls of bacteria, fungi, and yeast. Interestingly, the antimicrobial activity of AcMRJP4-15 was greater than that of AcMRJP4, demonstrating that the antimicrobial activity of AcMRJP4 was due in large part to the C-terminal. Our data suggest that AcMRJP4-15 can function as an effective antimicrobial agent.     

Assessment of the floral origin of honey via proteomic tools

The honey from chestnut, acacia, sunflower, eucalyptus and orange was analysed for its proteome content, in order to see if any plant proteins present would allow the proteo-typing of these different varieties. Since the total protein content turned out to be minute, 200g of each honey type were diluted to 1L and then added with ProteoMiner to enhance the visibility of the proteinaceous material. All bands visible in the SDS-PAGE profile of each type of honey were eluted, digested and identified by mass spectrometry in a LTQ-XL instrument. It turned out that all proteins identified (except one, the enzyme glyceraldehyde-3-phosphate dehydrogenase from Mesembryanthemum crystallinum) were not of plant origin but belonged to the Apis mellifera proteome. Among the total proteins identified (eight, but only seven as basic constituents of all types of honey) five belonged to the family of major royal jelly proteins 1-5, and were also the most abundant ones in any type of honey, together with α-glucosidase and defensin-1. It thus appears that honey has a proteome resembling the royal jelly proteome (but with considerably fewer species), except that its protein concentration is lower by three to four orders of magnitude as compared to royal jelly. Attempts at identifying additional plant (pollen, nectar) proteins via peptidome analysis were unsuccessful.     

Behavior and molecular physiology of nurses of worker and queen larvae in honey bees (Apis mellifera)

In a honey bee colony, worker bees rear a new queen by providing her with a larger cell in which to develop and a large amount of richer food (royal jelly). Royal jelly and worker jelly (fed to developing worker larvae) differ in terms of sugar, vitamin, protein and nucleotide composition. Here we examined whether workers attending queen and worker larvae are separate specialized sub-castes of the nurse bees. We collected nurse bees attending queen larvae (AQL) and worker larvae (AWL) and compared gene expression profiles of hypopharyngeal gland tissues, using Solexa/Illumina digital gene expression tag profiling (DGE). Significant differences in gene expression were found that included a disproportionate number of genes involved in glandular secretion and royal jelly synthesis. However behavioral observations showed that these were not two entirely distinct populations. Nurse workers were observed attending both worker larvae and queen larvae, and there was no evidence of a specialized group of workers that preferentially or exclusively attended developing queens. Nevertheless, AQL attended larvae more frequently compared to AWL, suggesting that nurses sampled attending queen larvae may have been the most active nurses. This study serves as another example of the relationship between differences in gene expression and behavioral specialisation in honey bees.     

Nephroprotective effect of bee honey and royal jelly against subchronic cisplatin toxicity in rats

Cisplatin is one of the most potent and effective chemotherapeutic agents. However, its antineoplastic use is limited due to its cumulative nephrotoxic side effects. Therefore, the present study was undertaken to examine the nephroprotective potential of dietary bee honey and royal jelly against subchronic cisplatin toxicity in rats. Male Wistar rats were randomly divided into controls, cisplatin-treated, bee honey-pretreated cisplatin-treated and royal jelly-pretreated cisplatin-treated groups. Bee honey and royal jelly were given orally at doses of 20 and 100 mg/kg, respectively. Subchronic toxicity was induced by cisplatin (1 mg/kg bw, ip), twice weekly for 10 weeks. Cisplatin treated animals revealed a significant increase in serum level of renal injury products (urea, creatinine and uric acid). Histopathologically, cisplatin produced pronounced tubulointerstitial injuries, upregulated the fibrogenic factors, α-smooth muscle actin (α-SMA) and transforming growth factor β1(TGF-β1), and downregulated the cell proliferation marker, bromodeoxyuridine (Brdu). Dietary bee honey and royal jelly normalized the elevated serum renal injury product biomarkers, improved the histopathologic changes, reduced the expression of α-SMA and TGF-β1 and increased the expression of Brdu. Therefore, it could be concluded that bee honey, and royal jelly could be used as dietary preventive natural products against subchronic cisplatin-induced renal injury.