Browsing by Subject "BIOSYNTHESIS"

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  • George, Jack; Tuomela, Tea; Kemppainen, Esko; Nurminen, Antti; Braun, Samuel; Yalgin, Cagri; Jacobs, Howard T. (2019)
    ABSTRACTThe Drosophila bang-sensitive mutant tko25t, manifesting a global deficiency in oxidative phosphorylation due to a mitochondrial protein synthesis defect, exhibits a pronounced delay in larval development. We previously identified a number of metabolic abnormalities in tko25t larvae, including elevated pyruvate and lactate, and found the larval gut to be a crucial tissue for the regulation of larval growth in the mutant. Here we established that expression of wild-type tko in any of several other tissues of tko25t also partially alleviates developmental delay. The effects appeared to be additive, whilst knockdown of tko in a variety of specific tissues phenocopied tko25t, producing developmental delay and bang-sensitivity. These findings imply the existence of a systemic signal regulating growth in response to mitochondrial dysfunction. Drugs and RNAi-targeted on pyruvate metabolism interacted with tko25t in ways that implicated pyruvate or one of its metabolic derivatives in playing a central role in generating such a signal. RNA-seq revealed that dietary pyruvate-induced changes in transcript representation were mostly non-coherent with those produced by tko25t or high-sugar, consistent with the idea that growth regulation operates primarily at the translational and/or metabolic level.
  • Fewer, David P.; Jokela, Jouni; Paukku, Eeva; Osterholm, Julia; Wahlsten, Matti; Permi, Perttu; Aitio, Olli; Rouhiainen, Leo; Gomez-Saez, Gonzalo V.; Sivonen, Kaarina (2013)
  • Olkkonen, Vesa M.; Koponen, Annika; Arora, Amita (2019)
    Oxysterol-binding protein (OSBP)-related proteins (ORPs) constitute a family of intracellular lipid-binding/transport proteins (LTPs) in eukaryotes. They typically have a modular structure comprising a lipid-binding domain and membrane targeting determinants, being thus suited for function at membrane contact sites. Among the mammalian ORPs, ORP2/OSBPL2 is the only member that only exists as a 'short' variant lacking a membrane-targeting pleckstrin homology domain. ORP2 is expressed ubiquitously and has been assigned a multitude of functions. Its OSBP-related domain binds cholesterol, oxysterols, and phosphoinositides, and its overexpression enhances cellular cholesterol efflux. Consistently, the latest observations suggest a function of ORP2 in cholesterol transport to the plasma membrane (PM) in exchange for phosphatidylinositol 4,5-bisphosphate (PI4,5P(2)), with significant impacts on the concentrations of PM cholesterol and PI4,5P(2). On the other hand, ORP2 localizes at the surface of cytoplasmic lipid droplets (LDs) and at endoplasmic-reticulum-LD contact sites, and its depletion modifies cellular triglyceride (TG) metabolism. Study in an adrenocortical cell line further suggested a function of ORP2 in the synthesis of steroid hormones. Our recent knock-out of ORP2 in human hepatoma cells revealed its function in hepatocellular PI3K/Akt signaling, glucose and triglyceride metabolism, as well as in actin cytoskeletal regulation, cell adhesion, migration and proliferation. ORP2 was shown to interact physically with F-actin regulators such as DIAPH1, ARHGAP12, SEPT9 and MLC12, as well as with IQGAP1 and the Cdc37-Hsp90 chaperone complex controlling the activity of Akt. Interestingly, mutations in OSBPL2 encoding ORP2 are associated with autosomal dominant non-syndromic hearing loss, and the protein was found to localize in cochlear hair cell stereocilia. The functions assigned to ORP2 suggest that this protein, in concert with other LTPs, controls the subcellular distribution of cholesterol in various cell types and steroid hormone synthesis in adrenocortical cells. However, it also impacts cellular TG and carbohydrate metabolism and F-actin-dependent functions, revealing a bewildering spectrum of activities.
  • Natsume, Masahiro; Nagagata, Asaho; Aittamaa, Marja; Okaniwa, Naoko; Somervuo, Panu; Fiedler, Hans-Peter; Kreuze, Jan F.; Rokka, Veli-Matti; Bang, Hans; Kawaide, Hiroshi; Valkonen, Jari P. T. (2018)
    Streptomyces spp. are a highly diverse group of bacteria most of which are soil-inhabiting saprophytes. A few are plant pathogens that produce a family of phytotoxins called thaxtomins and cause significant economic losses, e.g., by reducing the marketability of potato tubers (Solanum tuberosum). In northern Europe, S. scabies, S. turgidiscabies and S. europaeiscabiei are the most common plant pathogenic species. In this study, a Streptomyces strain isolated from a netted scab lesion on a tuber of potato cv. Bintje in northern Sweden was identified as S. turgidiscabies but was found to differ in the genomic region carrying genes required for thaxtomin biosynthesis. Our results showed that the strain did not produce thaxtomin but rather phytotoxin fridamycin E, which is an anthraquinone novel to plant pathogenic Streptomyces spp. Fridamycin E was shown to reduce or inhibit sprouting of potato microtubers in vitro. While fridamycin E is known to have antibiotic activity against Gram-positive bacteria, the inhibitory activity of fridamycin E on plant growth is a novel finding.
  • Seyfferth, Carolin; Wessels, Bernard A.; Vahala, Jorma; Kangasjarvi, Jaakko; Delhomme, Nicolas; Hvidsten, Torgeir R.; Tuominen, Hannele; Lundberg-Felten, Judith (2021)
    Secondary growth relies on precise and specialized transcriptional networks that determine cell division, differentiation, and maturation of xylem cells. We identified a novel role for the ethylene-induced Populus Ethylene Response Factor PtERF85 (Potri.015G023200) in balancing xylem cell expansion and secondary cell wall (SCW) formation in hybrid aspen (Populus tremula x tremuloides). Expression of PtERF85 is high in phloem and cambium cells and during the expansion of xylem cells, while it is low in maturing xylem tissue. Extending PtERF85 expression into SCW forming zones of woody tissues through ectopic expression reduced wood density and SCW thickness of xylem fibers but increased fiber diameter. Xylem transcriptomes from the transgenic trees revealed transcriptional induction of genes involved in cell expansion, translation, and growth. The expression of genes associated with plant vascular development and the biosynthesis of SCW chemical components such as xylan and lignin, was down-regulated in the transgenic trees. Our results suggest that PtERF85 activates genes related to xylem cell expansion, while preventing transcriptional activation of genes related to SCW formation. The importance of precise spatial expression of PtERF85 during wood development together with the observed phenotypes in response to ectopic PtERF85 expression suggests that PtERF85 contributes to the transition of fiber cells from elongation to secondary cell wall deposition.
  • Koivusalo, Antti; Pakarinen, Mikko; Gylling, Helena; Nissinen, Markku J. (2015)
    Background: Cholesterol metabolism may be involved in pediatric gallstone disease. We aimed to reveal cholesterol metabolites and phytosterols and their relation to stone composition of sterols in children having black pigment and cholesterol stones. Methods: We performed retrospective controlled clinical study, in which we examined parameters of cholesterol metabolism and liver function values in serum (n = 28) and gallstones (n = 46) of consecutively cholecystectomized children. Serum values of age-, body mass index-and sex-matched children (n = 82) and adult gallstones (n = 187) served as controls. Results: Surrogate markers of cholesterol synthesis in serum (squalene/cholesterol, cholestenol/cholesterol and lathosterol/cholesterol) were 26-52 % higher in both stone subclasses compared to controls (p <0.05 for all). Respectively, cholestanol/cholesterol and plant sterols campesterol/cholesterol and sitosterol/cholesterol (cholesterol absorption markers) had decreasing order in serum: black pigment stone group > controls > cholesterol stone group (p <0.05 for all). In black pigment stone group, stone cholestanol/cholesterol was associated with serum bile acids (r = 0.620, p = 0.018). In cholesterol stone group, surrogate markers of cholesterol synthesis in serum (e.g., lathosterol/cholesterol) inversely reflected those of absorption (r-range -0.633--0.706, p-range 0.036-0.015). In cholesterol stone group, serum and stone lathosterol/cholesterol and cholestanol/cholesterol were positively interrelated (r-range 0.727-0.847, p <0.05 for both). Conclusions: Gallstone subclasses shared enhanced cholesterol synthesis. Cholesterol stone children were low cholesterol absorbers with intact homeostasis of cholesterol metabolism. Black pigment stone group was characterized by deteriorated cholesterol metabolism, and accumulation of cholestanol, campesterol and sitosterol in serum and stones suggesting their participation in pathogenesis.
  • Jiang, Xiaojie; Keto-Timonen, Riikka; Skurnik, Mikael; Korkeala, Hannu (2019)
    Yersinia pseudotuberculosis is an important foodborne pathogen threatening modern food safety due to its ability to survive and grow at low temperatures. DEAD-box RNA helicase CsdA has been shown to play an important role in the low-temperature growth of psychrotrophic Y. pseudotuberculosis. A total of five DEAD-box RNA helicase genes (rhlB, csdA, rhlE, dbpA, srmB) have been identified in Y. pseudotuberculosis IP32953. However, their role in various stress conditions used in food production is unclear. We studied the involvement of the DEAD-box RNA helicase-encoding genes in the cold tolerance of Y. pseudotuberculosis IP32953 using quantitative real-time reverse transcription (RT-qPCR) and mutational analysis. Quantitative RT-PCR revealed that mRNA transcriptional levels of csdA, rhlE, dbpA and srmB were significantly higher after cold shock at 3 degrees C compared to non-shocked culture at 28 degrees C, suggesting the involvement of these four genes in cold shock response at the transcriptional level. The deletion of csdA ceased growth, while the deletion of dbpA or srmB significantly impaired growth at 3 degrees C, suggesting the requirement of these three genes in Y. pseudotuberculosis at low temperatures. Growth of each DEAD-box RNA helicase mutant was also investigated under pH, osmotic, ethanol and oxidative stress conditions. The five helicase-encoding genes did not play major roles in the growth of Y. pseudotuberculosis IP32953 under pH, osmotic, ethanol or oxidative stress.
  • Eronen-Rasimus, Eeva Liisa; Hultman, Jenni; Hai, T; Pessi, Igor S; Collins, Eric; Wright, S; Laine, Pia; Viitamäki, Sirja; Lyra, Christina; Thomas, David Neville; Golyshin, Peter; Luhtanen, Anne-Mari; Kuosa, Harri; Kaartokallio, Hermanni (2021)
    Poly-3-hydroxyalkanoic acids (PHAs) are bacterial storage polymers commonly used in bioplastic production. Halophilic bacteria are industrially interesting organisms, as their salinity tolerance and psychrophilic nature lowers sterility requirements and subsequent production costs. We investigated PHA synthesis in two bacterial strains, Halomonas sp. 363 and Paracoccus sp. 392, isolated from Southern Ocean sea ice and elucidated the related PHA biopolymer accumulation and composition with various approaches, such as transcriptomics, microscopy, and chromatography. We show that both bacterial strains produce PHAs at 4 degrees C when the availability of nitrogen and/or oxygen limited growth. The genome of Halomonas sp. 363 carries three phaC synthase genes and transcribes genes along three PHA pathways (I to III), whereas Paracoccus sp. 392 carries only one phaC gene and transcribes genes along one pathway (I). Thus, Halomonas sp. 363 has a versatile repertoire of phaC genes and pathways enabling production of both short- and medium-chain-length PHA products. IMPORTANCE Plastic pollution is one of the most topical threats to the health of the oceans and seas. One recognized way to alleviate the problem is to use degradable bioplastic materials in high-risk applications. PHA is a promising bioplastic material as it is nontoxic and fully produced and degraded by bacteria. Sea ice is an interesting environment for prospecting novel PHA-producing organisms, since traits advantageous to lower production costs, such as tolerance for high salinities and low temperatures, are common. We show that two sea-ice bacteria, Halomonas sp. 363 and Paracoccus sp. 392, are able to produce various types of PHA from inexpensive carbon sources. Halomonas sp. 363 is an especially interesting PHA-producing organism, since it has three different synthesis pathways to produce both short- and medium-chain-length PHAs.
  • Peret, Benjamin; Middleton, Alistair M.; French, Andrew P.; Larrieu, Antoine; Bishopp, Anthony; Njo, Maria; Wells, Darren M.; Porco, Silvana; Mellor, Nathan; Band, Leah R.; Casimiro, Ilda; Kleine-Vehn, Juergen; Vanneste, Steffen; Sairanen, Ilkka; Mallet, Romain; Sandberg, Goran; Ljung, Karin; Beeckman, Tom; Benkova, Eva; Friml, Jiri; Kramer, Eric; King, John R.; De Smet, Ive; Pridmore, Tony; Owen, Markus; Bennett, Malcolm J. (2013)
  • Kenyon, Johanna J.; Duda, Katarzyna A.; De Felice, Antonia; Cunneen, Monica M.; Molinaro, Antonio; Laitinen, Juha; Skurnik, Mikael; Holst, Otto; Reeves, Peter R.; De Castro, Cristina (2016)
    In Yersinia pseudotuberculosis complex, the O-antigen of LPS is used for the serological characterization of strains, and 21 serotypes have been identified to date. The O-antigen biosynthesis gene cluster and corresponding O-antigen structure have been described for 18, leaving O:8, O:13 and O:14 unresolved. In this study, two O:8 isolates were examined. The O-antigen gene cluster sequence of strain 151 was near identical to serotype O:4a, though a frame-shift mutation was found in ddhD, while No. 6 was different to 151 and carried the O:1b gene cluster. Structural analysis revealed that No. 6 produced a deeply truncated LPS, suggesting a mutation within the waaF gene. Both ddhD and waaF were cloned and expressed in 151 and No. 6 strains, respectively, and it appeared that expression of ddhD gene in strain 151 restored the O-antigen on LPS, while waaF in No. 6 resulted in an LPS truncated less severely but still without the O-antigen, suggesting that other mutations occurred in this strain. Thus, both O:8 isolates were found to be spontaneous O-antigen-negative mutants derived from other validated serotypes, and we propose to remove this serotype from the O-serotyping scheme, as the O:8 serological specificity is not based on the O-antigen.
  • Kalmbach, Lothar; Helariutta, Yrjö Eero (2019)
    Sieve pores of the sieve plates connect neighboring sieve elements to form the conducting sieve tubes of the phloem. Sieve pores are critical for phloem function. From the 1950s onwards, when electron microscopes became increasingly available, the study of their formation had been a pillar of phloem research. More recent work on sieve elements instead has largely focused on sieve tube hydraulics, phylogeny, and eco-physiology. Additionally, advanced molecular and genetic tools available for the model species Arabidopsis thaliana helped decipher several key regulatory mechanisms of early phloem development. Yet, the downstream differentiation processes which form the conductive sieve tube are still largely unknown, and our understanding of sieve pore formation has only moderately progressed. Here, we summarize our current knowledge on sieve pore formation and present relevant recent advances in related fields such as sieve element evolution, physiology, and plasmodesmata formation.
  • Mattila, Jaakko; Kokki, Krista; Hietakangas, Ville; Boutros, Michael (2018)
    The intestine is an organ with an exceptionally high rate of cell turnover, and perturbations in this process can lead to severe diseases such as cancer or intestinal atrophy. Nutrition has a profound impact on intestinal volume and cellular architecture. However, how intestinal homeostasis is maintained in fluctuating dietary conditions remains insufficiently understood. By utilizing the Drosophila midgut model, we reveal a novel stem cell intrinsic mechanism coupling cellular metabolism with stem cell extrinsic growth signal. Our results show that intestinal stem cells (ISCs) employ the hexosamine biosynthesis pathway (HBP) to monitor nutritional status. Elevated activity of HBP promotes Warburg effectlike metabolic reprogramming required for adjusting the ISC division rate according to nutrient content. Furthermore, HBP activity is an essential facilitator for insulin signaling-induced ISC proliferation. In conclusion, ISC intrinsic hexosamine synthesis regulates metabolic pathway activities and defines the stem cell responsiveness to niche-derived growth signals.
  • Toropainen, Sari; Malinen, Marjo; Kaikkonen, Sanna; Rytinki, Miia; Jääskeläinen, Tiina; Sahu, Biswajyoti; Janne, Olli A.; Palvimo, Jorma J. (2015)
  • Reigada, Inés; San-Martin-Galindo, Paola; Gilbert-Girard, Shella; Chiaro, Jacopo; Cerullo, Vincenzo; Savijoki, Kirsi; Nyman, Tuula A.; Fallarero, Adyary; Miettinen, Ilkka (2021)
    Bacterial biofilms are an important underlying cause for chronic infections. By switching into the biofilm state, bacteria can evade host defenses and withstand antibiotic chemotherapy. Despite the fact that biofilms at clinical and environmental settings are mostly composed of multiple microbial species, biofilm research has largely been focused on single-species biofilms. In this study, we investigated the interaction between two clinically relevant bacterial pathogens (Staphylococcus aureus and Pseudomonas aeruginosa) by label-free quantitative proteomics focusing on proteins associated with the bacterial cell surfaces (surfaceome) and proteins exported/released to the extracellular space (exoproteome). The changes observed in the surfaceome and exoproteome of P. aeruginosa pointed toward higher motility and lower pigment production when co-cultured with S. aureus. In S. aureus, lower abundances of proteins related to cell wall biosynthesis and cell division, suggesting increased persistence, were observed in the dual-species biofilm. Complementary phenotypic analyses confirmed the higher motility and the lower pigment production in P. aeruginosa when co-cultured with S. aureus. Higher antimicrobial tolerance associated with the co-culture setting was additionally observed in both species. To the best of our knowledge, this study is among the first systematic explorations providing insights into the dynamics of both the surfaceome and exoproteome of S. aureus and P. aeruginosa dual-species biofilms.
  • Lebeer, Sarah; Claes, Ingmar J. J.; Balog, Crina I. A.; Schoofs, Geert; Verhoeven, Tine L. A.; Nys, Kris; von Ossowski, Ingemar; de Vos, Willem M.; Tytgat, Hanne L. P.; Agostinis, Patrizia; Palva, Airi; Van Damme, Els J. M.; Deelder, Andre M.; De Keersmaecker, Sigrid C. J.; Wuhrer, Manfred; Vanderleyden, Jos (2012)
  • Filik, Karolina; Szermer-Olearnik, Bozena; Wernecki, Maciej; Happonen, Lotta J.; Pajunen, Maria I.; Nawaz, Ayesha; Qasim, Muhammad Suleman; Jun, Jin Woo; Mattinen, Laura; Skurnik, Mikael; Brzozowska, Ewa (2020)
    We report here the complete genome sequence and characterization ofYersiniabacteriophage vB_YenP_phi 80-18. phi 80-18 was isolated in 1991 using aY. enterocoliticaserotype O:8 strain 8081 as a host from a sewage sample in Turku, Finland, and based on its morphological and genomic features is classified as a podovirus. The genome is 42 kb in size and has 325 bp direct terminal repeats characteristic for podoviruses. The genome contains 57 predicted genes, all encoded in the forward strand, of which 29 showed no similarity to any known genes. Phage particle proteome analysis identified altogether 24 phage particle-associated proteins (PPAPs) including those identified as structural proteins such as major capsid, scaffolding and tail component proteins. In addition, also the DNA helicase, DNA ligase, DNA polymerase, 5 '-exonuclease, and the lytic glycosylase proteins were identified as PPAPs, suggesting that they might be injected together with the phage genome into the host cell to facilitate the take-over of the host metabolism. The phage-encoded RNA-polymerase and DNA-primase were not among the PPAPs. Promoter search predicted the presence of four phage and eleven host RNA polymerase -specific promoters in the genome, suggesting that early transcription of the phage is host RNA-polymerase dependent and that the phage RNA polymerase takes over later. The phage tolerates pH values between 2 and 12, and is stable at 50 degrees C but is inactivated at 60 degrees C. It grows slowly with a 50 min latent period and has apparently a low burst size. Electron microscopy revealed that the phage has a head diameter of about 60 nm, and a short tail of 20 nm. Whole-genome phylogenetic analysis confirmed that phi 80-18 belongs to theAutographivirinaesubfamily of thePodoviridaefamily, that it is 93.2% identical toYersiniaphage fHe-Yen3-01. Host range analysis showed that phi 80-18 can infect in addition toY. enterocoliticaserotype O:8 strains also strains of serotypes O:4, O:4,32, O:20 and O:21, the latter ones representing similar toY. enterocoliticaserotype O:8, the American pathogenic biotype 1B strains. In conclusion, the phage phi 80-18 is a promising candidate for the biocontrol of the American biotype 1BY. enterocolitica.
  • Deng, Xianbao; Elomaa, Paula; Nguyen, Cuong X.; Hytonen, Timo; Valkonen, Jari P. T.; Teeri, Teemu H. (2012)
  • Leskinen, Katarzyna; Pajunen, Maria I.; Vilanova, Miguel Vincente Gomez-Raya; Kiljunen, Saija; Nelson, Andrew; Smith, Darren; Skurnik, Mikael (2020)
    YerA41 is a Myoviridae bacteriophage that was originally isolated due its ability to infect Yersinia ruckeri bacteria, the causative agent of enteric redmouth disease of salmonid fish. Several attempts to determine its genomic DNA sequence using traditional and next generation sequencing technologies failed, indicating that the phage genome is modified in such a way that it is an unsuitable template for PCR amplification and for conventional sequencing. To determine the YerA41 genome sequence, we performed RNA-sequencing from phage-infected Y. ruckeri cells at different time points post-infection. The host-genome specific reads were subtracted and de novo assembly was performed on the remaining unaligned reads. This resulted in nine phage-specific scaffolds with a total length of 143 kb that shared only low level and scattered identity to known sequences deposited in DNA databases. Annotation of the sequences revealed 201 predicted genes, most of which found no homologs in the databases. Proteome studies identified altogether 63 phage particle-associated proteins. The RNA-sequencing data were used to characterize the transcriptional control of YerA41 and to investigate its impact on the bacterial gene expression. Overall, our results indicate that RNA-sequencing can be successfully used to obtain the genomic sequence of non-sequencable phages, providing simultaneous information about the phage–host interactions during the process of infection.