Similar articles

Abstract: Royal jelly (RJ), secreted by the hypopharyngeal and mandibular glands of young worker bees, is rich in proteins, 80%‒90% of which are major royal jelly proteins (MRJPs). While MRJPs from RJ have been shown to exhibit specific biological functions and are also expressed in neuronal cells, their roles in vivo remain poorly understood. The aim of this study was to elucidate the functional roles of individual MRJPs (MRJP1‒9) in vivo by ectopically expressing them in Drosophila neurons in a binary expression system using fluorescent proteins as controls. Transcriptome sequencing revealed that although MRJP1‒9 share similar tertiary structures, their overexpression affects distinct gene sets. MRJP1, MRJP2, MRJP3, MRJP5, and MRJP7 induced more differentially expressed genes (DEGs), while MRJP4, MRJP6, MRJP8, and MRJP9 induced fewer such genes. Weighted gene co-expression network analysis (WGCNA) and gene set enrichment analysis (GSEA) revealed that MRJP1, MRJP2, MRJP3, MRJP5, and MRJP7 regulated overlapping gene sets, including an estradiol-responsive set, and activated cell proliferation pathways. MRJP6 lacked any significant gene set enrichment, while MRJP8 and MRJP9 modulated similar sets. Notably, the neuron-specific overexpression of MRJP1, MRJP2, MRJP3, and MRJP5 in Drosophila showed activated cell proliferation-related pathways and increased body sizes, highlighting their functional diversity and context-dependent effects. These findings expand our understanding of the functional roles of MRJPs and provide a foundation for further exploring their biological significance in honeybees and beyond.

果蝇模型系统性解析蜂王浆主蛋白在神经细胞中的功能异质性

[1]AbramsonJ, AdlerJ, DungerJ, et al., 2024. Accurate structure prediction of biomolecular interactions with AlphaFold 3. Nature, 630(8016):493-500.

[2]Abu-SerieMM, HabashyNH, 2019. Two purified proteins from royal jelly with in vitro dual anti-hepatic damage potency: major royal jelly protein 2 and its novel isoform X1. Int J Biol Macromol, 128:782-795.

[3]AlbertŠ, SpaetheJ, GrübelK, et al., 2014. Royal jelly-like protein localization reveals differences in hypopharyngeal glands buildup and conserved expression pattern in brains of bumblebees and honeybees. Biol Open, 3(4):281-288.

[4]BílikováK, ŠimúthJ, 2010. New criterion for evaluation of honey: quantification of royal jelly protein apalbumin 1 in honey by ELISA. J Agric Food Chem, 58(15):8776-8781.

[5]BílikováK, MirgorodskayáE, BukovskaG, et al., 2009. Towards functional proteomics of minority component of honeybee royal jelly: the effect of post-translational modifications on the antimicrobial activity of apalbumin2. Proteomics, 9(8):2131-2138.

[6]ButtstedtA, MoritzRF, ErlerS, 2013. More than royal food‒Major royal jelly protein genes in sexuals and workers of the honeybee Apis mellifera. Front Zool, 10:72.

[7]ButtstedtA, IhlingCH, PietzschM, et al., 2016. Royalactin is not a royal making of a queen. Nature, 537(7621):E10-E12.

[8]ChakrabartiP, SagiliRR, 2020. Changes in honey bee head proteome in response to dietary 24-methylenecholesterol. Insects, 11(11):743.

[9]CollazoN, CarpenaM, Nuñez-EstevezB, et al., 2021. Health promoting properties of bee royal jelly: food of the queens. Nutrients, 13(2):543.

[10]ConnahsH, TanEJ, TerYT, et al., 2022. The yellow gene regulates behavioural plasticity by repressing male courtship in Bicyclus anynana butterflies. Proc Roy Soc B Biol Sci, 289(1972):20212665.

[11]DetienneG, de HaesW, ErnstUR, et al., 2014. Royalactin extends lifespan of Caenorhabditis elegans through epidermal growth factor signaling. Exp Gerontol, 60:129-135.

[12]DobritzschD, AumerD, FuszardM, et al., 2019. The rise and fall of major royal jelly proteins during a honeybee (Apis mellifera) workers’ life. Ecol Evol, 9(15):8771-8782.

[13]DrapeauMD, RadovicA, WittkoppPJ, et al., 2003. A gene necessary for normal male courtship, yellow, acts downstream of fruitless in the Drosophila melanogaster larval brain. J Neurobiol, 55(1):53-72.

[14]DrapeauMD, AlbertS, KucharskiR, et al., 2006. Evolution of the Yellow/Major Royal Jelly Protein family and the emergence of social behavior in honey bees. Genome Res, 16(11):1385-1394.

[15]FanP, HanB, FengM, et al., 2016. Functional and proteomic investigations reveal major royal jelly protein 1 associated with anti-hypertension activity in mouse vascular smooth muscle cells. Sci Rep, 6:30230.

[16]FangY, FengM, MaC, et al., 2023. Major royal jelly proteins influence the neurobiological regulation of the division of labor among honey bee workers. Int J Biol Macromol, 225:848-860.

[17]GarciaL, Saraiva GarciaCH, CalábriaLK, et al., 2009. Proteomic analysis of honey bee brain upon ontogenetic and behavioral development. J Proteome Res, 8(3):1464-1473.

[18]GuoH, KouzumaY, YonekuraM, 2009. Structures and properties of antioxidative peptides derived from royal jelly protein. Food Chem, 113(1):238-245.

[19]HayashiT, TakamatsuN, NakashimaT, et al., 2011. Immunological characterization of honey proteins and identification of MRJP 1 as an IgE-binding protein. Biosci Biotechnol Biochem, 75(3):556-560.

[20]HinauxH, BachemK, BattistaraM, et al., 2018. Revisiting the developmental and cellular role of the pigmentation gene yellow in Drosophila using a tagged allele. Dev Biol, 438(2):111-123.

[21]HojoM, KagamiT, SasakiT, et al., 2010. Reduced expression of major royal jelly protein 1 gene in the mushroom bodies of worker honeybees with reduced learning ability. Apidologie, 41(2):194-202.

[22]KamakuraM, 2011. Royalactin induces queen differentiation in honeybees. Nature, 473(7348):478-483.

[23]KamakuraM, SakakiT, 2006. A hypopharyngeal gland protein of the worker honeybee Apis mellifera L. enhances proliferation of primary-cultured rat hepatocytes and suppresses apoptosis in the absence of serum. Protein Expr Purif, 45(2):307-314.

[24]KamakuraM, SuenobuN, FukushimaM, 2001. Fifty-seven-kDa protein in royal jelly enhances proliferation of primary cultured rat hepatocytes and increases albumin production in the absence of serum. Biochem Biophys Res Commun, 282(4):865-874.

[25]KimBY, JinBR, 2019. Antimicrobial activity of the C-terminal of the major royal jelly protein 4 in a honeybee (Apis cerana). J Asia-Pac Entomol, 22(2):561-564.

[26]KimuraY, NagaiH, MiyamotoM, et al., 2010. Identification of a royal jelly glycoprotein that carries unique complex-type N-glycans harboring the T-antigen (Galβ1-3GalNAc) unit. Biosci Biotechnol Biochem, 74(10):2148-2150.

[27]KumarS, StecherG, LiM, et al., 2018. MEGA X: molecular evolutionary genetics analysis across computing platforms. Mol Biol Evol, 35(6):1547-1549.

[28]KupkeJ, SpaetheJ, MuellerMJ, et al., 2012. Molecular and biochemical characterization of the major royal jelly protein in bumblebees suggest a non-nutritive function. Insect Biochem Mol Biol, 42(9):647-654.

[29]LangfelderP, HorvathS, 2008. WGCNA: an R package for weighted correlation network analysis. BMC Bioinformatics, 9:559.

[30]LiberzonA, BirgerC, ThorvaldsdóttirH, et al., 2015. The Molecular Signatures Database hallmark gene set collection. Cell Syst, 1(6):417-425.

[31]LinY, ShaoQQ, ZhangM, et al., 2019. Royal jelly-derived proteins enhance proliferation and migration of human epidermal keratinocytes in an in vitro scratch wound model. BMC Complement Altern Med, 19:175.

[32]MajtánJ, KováčováE, BílikováK, et al., 2006. The immunostimulatory effect of the recombinant apalbumin 1‒major honeybee royal jelly protein‒on TNFα release. Int Immunopharmacol, 6(2):269-278.

[33]MaoriE, NavarroIC, BoncristianiH, et al., 2019. A secreted RNA binding protein forms RNA-stabilizing granules in the honeybee royal jelly. Mol Cell, 74(3):598-608.e6.

[34]MasseyJH, ChungD, SiwanowiczI, et al., 2019. The yellow gene influences Drosophila male mating success through sex comb melanization. eLife, 8:e49388.

[35]MatsuiT, YukiyoshiA, DoiS, et al., 2002. Gastrointestinal enzyme production of bioactive peptides from royal jelly protein and their antihypertensive ability in SHR. J Nutr Biochem, 13(2):80-86.

[36]MoothaVK, LindgrenCM, ErikssonKF, et al., 2003. PGC-1αresponsive genes involved in oxidative phosphorylation are coordinately downregulated in human diabetes. Nat Genet, 34(3):267-273.

[37]MoriyamaT, ItoA, OmoteS, et al., 2015. Heat resistant characteristics of Major Royal Jelly Protein 1 (MRJP1) oligomer. PLOS One, 10(5):e0119169.

[38]MureşanCI, DezmireanDS, MarcBD, et al., 2022. Biological properties and activities of major royal jelly proteins and their derived peptides. J Funct Foods, 98:105286.

[39]NagaiT, InoueR, 2004. Preparation and the functional properties of water extract and alkaline extract of royal jelly. Food Chem, 84(2):181-186.

[40]OkamotoI, TaniguchiY, KunikataT, et al., 2003. Major royal jelly protein 3 modulates immune responses in vitro and in vivo. Life Sci, 73(16):2029-2045.

[41]ParkMJ, KimBY, DengYJ, et al., 2020. Antioxidant capacity of major royal jelly proteins of honeybee (Apis mellifera) royal jelly. J Asia-Pac Entomol, 23(2):445-448.

[42]PeixotoLG, CalábriaLK, GarciaL, et al., 2009. Identification of major royal jelly proteins in the brain of the honeybee Apis mellifera. J Insect Physiol, 55(8):671-677.

[43]RadovicA, WittkoppPJ, LongAD, et al., 2002. Immunohistochemical colocalization of Yellow and male-specific fruitless in Drosophila melanogaster neuroblasts. Biochem Biophys Res Commun, 293(4):1262-1264.

[44]SchmitzováJ, KlaudinyJ, AlbertŠ, et al., 1998. A family of major royal jelly proteins of the honeybee Apis mellifera L. Cell Mol Life Sci, 54(9):1020-1030.

[45]ShenLR, ZhangWG, JinF, et al., 2010. Expression of recombinant AccMRJP1 protein from royal jelly of Chinese honeybee in Pichia pastoris and its proliferation activity in an insect cell line. J Agric Food Chem, 58(16):9190-9197.

[46]SubramanianA, TamayoP, MoothaVK, et al., 2005. Gene set enrichment analysis: a knowledge-based approach for interpreting genome-wide expression profiles. Proc Natl Acad Sci USA, 102(43):15545-15550.

[47]TamuraS, AmanoS, KonoT, et al., 2009. Molecular characteristics and physiological functions of major royal jelly protein 1 oligomer. Proteomics, 9(24):5534-5543.

[48]TianWL, LiM, GuoHY, et al., 2018. Architecture of the native major royal jelly protein 1 oligomer. Nat Commun, 9:3373.

[49]UverskyVN, AlbarAH, KhanRH, et al., 2021. Multifunctionality and intrinsic disorder of royal jelly proteome. Proteomics, 21(6):2000237.

[50]VezeteuTV, BobişO, MoritzRFA, et al., 2017. Food to some, poison to others - honeybee royal jelly and its growth inhibiting effect on European Foulbrood bacteria. Microbiologyopen, 6(1):e00397.

[51]WanDC, MorganSL, SpencleyAL, et al., 2018. Honey bee Royalactin unlocks conserved pluripotency pathway in mammals. Nat Commun, 9:5078.

[52]WatanabeK, ShinmotoH, KoboriM, et al., 1998. Stimulation of cell growth in the U-937 human myeloid cell line by honey royal jelly protein. Cytotechnology, 26(1):23-27.

[53]WenDJ, XieJY, YuanY, et al., 2024. The endogenous antioxidant ability of royal jelly in Drosophila is independent of Keap1/Nrf2 by activating oxidoreductase activity. Insect Sci, 31(2):503-523.

[54]XinXX, ChenY, ChenD, et al., 2016. Supplementation with major royal-jelly proteins increases lifespan, feeding, and fecundity in Drosophila. J Agric Food Chem, 64(29):5803-5812.

[55]ZhangL, HanB, LiRL, et al., 2014. Comprehensive identification of novel proteins and N-glycosylation sites in royal jelly. BMC Genomics, 15:135.

[56]ZhangXQ, YuY, SunP, et al., 2019. Royal jelly peptides: potential inhibitors of β-secretase in N2a/APP695swe cells. Sci Rep, 9:168.

[57]ZhangZJ, MuXH, CaoQN, et al., 2022. Honeybee gut Lactobacillus modulates host learning and memory behaviors via regulating tryptophan metabolism. Nat Commun, 13:2037.