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Visualizing trypanosomes in a vertebrate host reveals novel swimming behaviours, adaptations and attachment mechanisms
Dóró, Éva ; Jacobs, Sem H. ; Hammond, Ffion R. ; Schipper, Henk ; Pieters, Remco P.M. ; Carrington, Mark ; Wiegertjes, Geert F. ; Forlenza, Maria - \ 2019
eLife 8 (2019). - ISSN 2050-084X
host-pathogen interaction - infectious disease - microbiology - swimming behavior - Trypanosoma carassii - zebrafish
Trypanosomes are important disease agents of humans, livestock and cold-blooded species, including fish. The cellular morphology of trypanosomes is central to their motility, adaptation to the host's environments and pathogenesis. However, visualizing the behaviour of trypanosomes resident in a live vertebrate host has remained unexplored. In this study, we describe an infection model of zebrafish (Danio rerio) with Trypanosoma carassii. By combining high spatio-temporal resolution microscopy with the transparency of live zebrafish, we describe in detail the swimming behaviour of trypanosomes in blood and tissues of a vertebrate host. Besides the conventional tumbling and directional swimming, T. carassii can change direction through a 'whip-like' motion or by swimming backward. Further, the posterior end can act as an anchoring site in vivo. To our knowledge, this is the first report of a vertebrate infection model that allows detailed imaging of trypanosome swimming behaviour in vivo in a natural host environment.
Impulsive sounds change European seabass swimming patterns: influence of pulse repetition interval
Neo, Y.Y. ; Ufkes, E. ; Kastelein, R.A. ; Winter, H.V. ; Cate, C. ten; Slabbekoorn, H. - \ 2015
Marine Pollution Bulletin 97 (2015)1-2. - ISSN 0025-326X - p. 111 - 117.
startle-response - trawling vessel - fish - habituation - behavior - noise - variability - avoidance - zebrafish - recovery
Seismic shootings and offshore pile-driving are regularly performed, emitting significant amounts of noise that may negatively affect fish behaviour. The pulse repetition interval (PRI) of these impulsive sounds may vary considerably and influence the behavioural impact and recovery. Here, we tested the effect of four PRIs (0.5–4.0 s) on European seabass swimming patterns in an outdoor basin. At the onset of the sound exposures, the fish swam faster and dived deeper in tighter shoals. PRI affected the immediate and delayed behavioural changes but not the recovery time. Our study highlights that (1) the behavioural changes of captive European seabass were consistent with previous indoor and outdoor studies; (2) PRI could influence behavioural impact differentially, which may have management implications; (3) some acoustic metrics, e.g. SELcum, may have limited predictive power to assess the strength of behavioural impacts of noise. Noise impact assessments need to consider the contribution of sound temporal structure.
Ccbe1 regulates Vegfc-mediated induction of Vegfr3 signaling during embryonic lymphangiogenesis
Guen, L. ; Karpanen, T. ; Schulte, D. ; Harris, N.C. ; Koltowska, K. ; Roukens, G. ; Bower, N.I. ; Impel, A. van; Stacker, S.A. ; Achen, M.G. ; Schulte-Merker, S. ; Hogan, B.M. - \ 2014
Development 141 (2014). - ISSN 0950-1991 - p. 1239 - 1249.
growth-factor receptor-3 - lymphatic endothelial-cells - vascular development - factor-c - missense mutations - primary lymphedema - network formation - transgenic mice - in-vivo - zebrafish
The VEGFC/VEGFR3 signaling pathway is essential for lymphangiogenesis (the formation of lymphatic vessels from pre-existing vasculature) during embryonic development, tissue regeneration and tumor progression. The recently identified secreted protein CCBE1 is indispensible for lymphangiogenesis during development. The role of CCBE1 orthologs is highly conserved in zebrafish, mice and humans with mutations in CCBE1 causing generalized lymphatic dysplasia and lymphedema (Hennekam syndrome). To date, the mechanism by which CCBE1 acts remains unknown. Here, we find that ccbe1 genetically interacts with both vegfc and vegfr3 in zebrafish. In the embryo, phenotypes driven by increased Vegfc are suppressed in the absence of Ccbe1, and Vegfc-driven sprouting is enhanced by local Ccbe1 overexpression. Moreover, Vegfc- and Vegfr3-dependent Erk signaling is impaired in the absence of Ccbe1. Finally, CCBE1 is capable of upregulating the levels of fully processed, mature VEGFC in vitro and the overexpression of mature VEGFC rescues ccbe1 loss-of-function phenotypes in zebrafish. Taken together, these data identify Ccbe1 as a crucial component of the Vegfc/Vegfr3 pathway in the embryo.
Atypical E2fs Control Lymphangiogenesis through Transcriptional Regulation of Ccbe1 and Flt4
Weijts, B.G.M.W. ; Impel, A. van; Schulte-Merker, S. ; Bruin, A. de - \ 2013
PLoS ONE 8 (2013)9. - ISSN 1932-6203
endothelial-growth-factor - large gene lists - vascular development - plc-gamma - factor-c - in-vivo - cells - zebrafish - activation - angiogenesis
Lymphatic vessels are derived from venous endothelial cells and their formation is governed by the Vascular endothelial growth factor C (VegfC)/Vegf receptor 3 (Vegfr3; Flt4) signaling pathway. Recent studies show that Collagen and Calcium Binding EGF domains 1 protein (Ccbe1) enhances VegfC-dependent lymphangiogenesis. Both Ccbe1 and Flt4 have been shown to be indispensable for lymphangiogenesis. However, how these essential players are transcriptionally regulated remains poorly understood. In the case of angiogenesis, atypical E2fs (E2f7 and E2f8) however have been recently shown to function as transcriptional activators for VegfA. Using a genome-wide approach we here identified both CCBE1 and FLT4 as direct targets of atypical E2Fs. E2F7/8 directly bind and stimulate the CCBE1 promoter, while recruitment of E2F7/8 inhibits the FLT4 promoter. Importantly, inactivation of e2f7/8 in zebrafish impaired venous sprouting and lymphangiogenesis with reduced ccbe1 expression and increased flt4 expression. Remarkably, over-expression of e2f7/8 rescued Ccbe1- and Flt4-dependent lymphangiogenesis phenotypes. Together these results identified E2f7/8 as novel in vivo transcriptional regulators of Ccbe1 and Flt4, both essential genes for venous sprouting and lymphangiogenesis.
Inflammatory Chemokines Direct and Restrict Leukocyte Migration within Live Tissues as Glycan-Bound Gradients
Sarris, M. ; Masson, J.B. ; Maurin, D. ; Aa, L.M. van der; Boudinot, P. ; Lortat-Jacob, H. ; Herbomel, P. - \ 2012
Current Biology 22 (2012)24. - ISSN 0960-9822 - p. 2375 - 2382.
cd8(+) t-cells - heparan-sulfate - neutrophil migration - dendritic cells - in-vivo - zebrafish - chemotaxis - motility - trafficking - attraction
Chemokines are essential in many cell migration processes, including the recruitment of leukocytes to sites of infection [1-3]. In the latter context, chemokines promote leukocyte extravasation into the relevant tissue through a well-studied cascade of events [4-9]. It is widely believed that chemokines further guide leukocytes within tissues via chemotaxis, the directed migration along gradients of soluble ligands. However, the basic mechanism of chemokine action within tissues has yet to be formally addressed in vivo. We identified a chemokine (zCxcl8) that recruits zebrafish neutrophils to infection loci and analyzed its function directly within interstitial tissues of living larvae. Using noninvasive imaging and a controlled cellular source of zCxcl8, we found that zCxcl8 guides neutrophils in a 2-fold manner: by biasing cell speed according to direction (orthotaxis) and by restricting cell motility near the source. We further show that zCxcl8 establishes tissue-bound gradients in vivo by binding to heparan sulfate proteoglycans (HSPGs). Inhibition of this interaction compromised both directional guidance and restriction of neutrophil motility. Thus, by interacting with extracellular HSPGs, chemokines establish robust surface-bound (haptotactic) gradients that mediate both recruitment and retention of leukocytes at sites of infection.
E2F7 and E2F8 promote angiogenesis through transcriptional activation of VEGFA in cooperation with HIF1
Weijts, B.G.M.W. ; Bakker, W.J. ; Cornelissen, P.W.A. ; Liang, K. ; Schaftenaar, F.H. ; Westendorp, B. ; Wolf, C.A.C.M.T. de; Paciejewska, M. ; Scheele, C.L.G.J. ; Kent, L. ; Leone, G. ; Schulte-Merker, S. ; Bruin, A. de - \ 2012
The EMBO Journal 31 (2012)19. - ISSN 0261-4189 - p. 3871 - 3884.
hypoxia-inducible factor-1 - growth-factor gene - embryonic-development - vascular development - o-2 homeostasis - factor 1-alpha - zebrafish - proliferation - cancer - hif-1-alpha
The E2F family of transcription factors plays an important role in controlling cell-cycle progression. While this is their best-known function, we report here novel functions for the newest members of the E2F family, E2F7 and E2F8 (E2F7/8). We show that simultaneous deletion of E2F7/8 in zebrafish and mice leads to severe vascular defects during embryonic development. Using a panel of transgenic zebrafish with fluorescent-labelled blood vessels, we demonstrate that E2F7/8 are essential for proper formation of blood vessels. Despite their classification as transcriptional repressors, we provide evidence for a molecular mechanism through which E2F7/8 activate the transcription of the vascular endothelial growth factor A (VEGFA), a key factor in guiding angiogenesis. We show that E2F7/8 directly bind and stimulate the VEGFA promoter independent of canonical E2F binding elements. Instead, E2F7/8 form a transcriptional complex with the hypoxia inducible factor 1 (HIF1) to stimulate VEGFA promoter activity. These results uncover an unexpected link between E2F7/8 and the HIF1-VEGFA pathway providing a molecular mechanism by which E2F7/8 control angiogenesis.
Body dynamics and hydrodynamics of swimming larvae: a computational study
Li, G. ; Müller, U.K. ; Leeuwen, J.L. van; Liu, H. - \ 2012
Journal of Experimental Biology 215 (2012). - ISSN 0022-0949 - p. 4015 - 4033.
inertial flow regimes - escape response - kinematics - locomotion - wake - performance - waves - morphology - zebrafish - fields
To understand the mechanics of fish swimming, we need to know the forces exerted by the fluid and how these forces affect the motion of the fish. To this end, we developed a 3-D computational approach that integrates hydrodynamics and body dynamics. This study quantifies the flow around a swimming zebrafish (Danio rerio) larva. We used morphological and kinematics data from actual fish larvae aged 3 and 5 days post fertilization as input for a computational model that predicted free-swimming dynamics from prescribed changes in body shape. We simulated cyclic swimming and a spontaneous C-start. A rigorous comparison with 2-D particle image velocimetry and kinematics data revealed that the computational model accurately predicted the motion of the fish's centre of mass as well as the spatial and temporal characteristics of the flow. The distribution of pressure and shear forces along the body showed that thrust is mainly produced in the posterior half of the body. We also explored the effect of the body wave amplitude on swimming performance by considering wave amplitudes that were up to 40% larger or smaller than the experimentally observed value. Increasing the body wave amplitude increased forward swimming speed from 7 to 21 body lengths per second, which is consistent with experimental observations. The model also predicted a non-linear increase in propulsive efficiency from 0.22 to 0.32 while the required mechanical power quadrupled. The efficiency increase was only minor for wave amplitudes above the experimental reference value, whereas the cost of transport rose significantly.
Structure and function of the median finfold in larval teleosts
Boogaart, J.G.M. van den; Muller, M. ; Osse, J.W.M. - \ 2012
Journal of Experimental Biology 215 (2012). - ISSN 0022-0949 - p. 2359 - 2368.
fish larvae - allometric growth - engraulis-mordax - dermal skeleton - danio-rerio - body-size - zebrafish - fin - behavior - hydrodynamics
This paper offers a structural and mechanical analysis of the median finfold in larval teleosts. The median finfold is strengthened by bundles of collagen fibres, known as actinotrichia. We demonstrate that these structures contribute to increase the mass of backward accelerated water during swimming. The amount, dimensions, orientation and growth of actinotrichia were measured at various locations along the finfold in several developmental stages of common carp (Cyprinus carpio) and zebrafish (Danio rerio). Actinotrichia morphology, using light microscopy (e.g. diameter, orientation) and electron microscopy (which revealed their anchoring at proximal and distal ends), correlated with expected lateral forces exerted on the water during swimming. An analytical model is proposed that predicts the extent of camber from the oblique arrangement of the actinotrichia and curvature of the body. Camber of the finfold during swimming was measured from high-speed video recordings and used to evaluate the model predictions. Based on structural requirements for swimming and strain limits for collagen, the model also predicts optimal orientations of actinotrichia. Experimental data confirm the predictions of the model.
Trpv5/6 is vital for epithelial calcium uptake and bone formation
Vanoevelen, J. ; Janssens, A. ; Huitema, L.F.A. ; Hammond, C.L. ; Metz, J.R. ; Flik, G. ; Voets, T. ; Schulte-Merker, S. - \ 2011
FASEB Journal 25 (2011)9. - ISSN 0892-6638 - p. 3197 - 3207.
ca2+ channel - in-vivo - stannius corpuscles - cation channels - stanniocalcin 1 - danio-rerio - absorption - zebrafish - transport - receptor
Calcium is an essential ion serving a multitude of physiological roles. Aside from its role as a second messenger, it is an essential component of the vertebrate bone matrix. Efficient uptake and storage of calcium are therefore indispensable for all vertebrates. Transient receptor potential family, vanilloid type (TRPV)5 and TRPV6 channels are known players in transcellular calcium uptake, but the exact contribution of this pathway is unclear. We used forward genetic screening in zebrafish (Danio rerio) to identify genes essential in bone formation and identified a lethal zebrafish mutant (matt-und-schlapp) with severe defects in bone formation, including lack of ossification of the vertebral column and craniofacial structures. Mutant embryos show a 68% reduction in calcium content, and systemic calcium homeostasis is disturbed when compared with siblings. The phenotype can be partially rescued by increasing ambient calcium levels to 25 mM. We identified the mutation as a loss-of-function mutation in the single orthologue of TRPV5 and 6, trpv5/6. Expression in HEK293 cells showed that Trpv5/6 is a calcium-selective channel capable of inward calcium transport at physiological concentrations whereas the mutant channel is not. Taken together, this study provides both genetic and functional evidence that transcellular epithelial calcium uptake is vital to sustain life and enable bone formation.—Vanoevelen, J., Janssens, A., Huitema, L. F. A., Hammond, C. L., Metz, J. R., Flik, G., Voets, T., Schulte-Merker, S. Trpv5/6 is vital for epithelial calcium uptake and bone formation
Implementation of two high through-put techniques in a novel application: detecting point mutations in large EMS mutated plant populations
Gady, A.L.F. ; Hermans, F.W.K. ; Wal, M.H.B.J. van de; Loo, E.N. van; Visser, R.G.F. ; Bachem, C.W.B. - \ 2009
Plant Methods 5 (2009). - ISSN 1746-4811 - 14 p.
sensitive capillary-electrophoresis - chemically-induced mutations - resolution melting analysis - arabidopsis-thaliana - gene mutation - in-silico - discovery - tomato - zebrafish - disease
Background - The establishment of mutant populations together with the strategies for targeted mutation detection has been applied successfully to a large number of organisms including many species in the plant kingdom. Considerable efforts have been invested into research on tomato as a model for berry-fruit plants. With the progress of the tomato sequencing project, reverse genetics becomes an obvious and achievable goal. Results - Here we describe the treatment of Solanum lycopersicum seeds with 1% EMS and the development of a new mutated tomato population. To increase targeted mutant detection throughput an automated seed DNA extraction has been combined with novel mutation detection platforms for TILLING in plants. We have adapted two techniques used in human genetic diagnostics: Conformation Sensitive Capillary Electrophoresis (CSCE) and High Resolution DNA Melting Analysis (HRM) to mutation screening in DNA pools. Classical TILLING involves critical and time consuming steps such as endonuclease digestion reactions and gel electrophoresis runs. Using CSCE or HRM, the only step required is a simple PCR before either capillary electrophoresis or DNA melting curve analysis. Here we describe the development of a mutant tomato population, the setting up of two polymorphism detection platforms for plants and the results of the first screens as mutation density in the populations and estimation of the false-positives rate when using HRM to screen DNA pools. Conclusion - These results demonstrate that CSCE and HRM are fast, affordable and sensitive techniques for mutation detection in DNA pools and therefore allow the rapid identification of new allelic variants in a mutant population. Results from the first screens indicate that the mutagen treatment has been effective with an average mutation detection rate per diploid genome of 1.36 mutation/kb/1000 lines
Flow patterns of larval fish: undulatory swimming in the intermediate flow regime
Müller, U.K. ; Boogaart, J.G.M. van den; Leeuwen, J.L. van - \ 2008
Journal of Experimental Biology 211 (2008)2. - ISSN 0022-0949 - p. 196 - 205.
locomotor function - rainbow-trout - vortex wake - dorsal fin - hydrodynamics - kinematics - zebrafish - forces - generation - morphology
Fish larvae, like many adult fish, swim by undulating their body. However, their body size and swimming speeds put them in the intermediate flow regime, where viscous and inertial forces both play an important role in the interaction between fish and water. To study the influence of the relatively high viscous forces compared with adult fish, we mapped the flow around swimming zebrafish (Danio rerio) larvae using two-dimensional digital particle image velocimetry (2D-DPIV) in the horizontal and transverse plane of the fish. Fish larvae initiate a swimming bout by bending their body into a C shape. During this initial tail-beat cycle, larvae shed two vortex pairs in the horizontal plane of their wake, one during the preparatory and one during the subsequent propulsive stroke. When they swim `cyclically' (mean swimming speed does not change significantly between tail beats), fish larvae generate a wide drag wake along their head and anterior body. The flow along the posterior body is dominated by the undulating body movements that cause jet flows into the concave bends of the body wave. Patches of elevated vorticity form around the jets, and travel posteriorly along with the body wave, until they are ultimately shed at the tail near the moment of stroke reversal. Behind the larva, two vortex pairs are formed per tail-beat cycle (the tail beating once left-to-right and then right-to-left) in the horizontal plane of the larval wake. By combining transverse and horizontal cross sections of the wake, we inferred that the wake behind a cyclically swimming zebrafish larva contains two diverging rows of vortex rings to the left and right of the mean path of motion, resembling the wake of steadily swimming adult eels. When the fish larva slows down at the end of a swimming bout, it gradually reduces its tail-beat frequency and amplitude, while the separated boundary layer and drag wake of the anterior body extend posteriorly to envelope the entire larva. This drag wake is considerably wider than the larval body. The effects of the intermediate flow regime manifest as a thick boundary layer and in the quick dying-off of the larval wake within less than half a second
Ontogeny of the common carp (Cyprinus carpio L.) innate immune system
Huttenhuis, B.T. ; Taverne-Thiele, J.J. ; Grou, C.P.O. ; Bergsma, J. ; Saeij, J.P.J. ; Nakayasu, C. ; Rombout, J.H.W.M. - \ 2006
Developmental and Comparative Immunology 30 (2006)6. - ISSN 0145-305X - p. 557 - 574.
complement component c3 - functional-characterization - nitric-oxide - expression analysis - molecular-cloning - early macrophages - zebrafish - fish - hematopoiesis - interleukin-1-beta
The ontogeny of the teleost innate immune system was studied in carp using cellular, histological and quantitative molecular techniques. Carp myeloid cells first appeared ventro-lateral of the aorta at 2 days post fertilization (the start of hatching), and subsequently around the sinuses of the vena cardinalis (or posterior blood islet), head kidney and trunk kidney. In addition, the hematopoietic tissue around the sinuses of the vena cardinalis transformed into that of the trunk kidney, which is the first description of the ontogeny of the trunk kidney hematopoietic tissue in teleosts. The mAb's used in this study reacted with carp myeloid surface molecules that are already transcribed and processed from the first appearance of myeloid cells, and thus serve a significant role in unraveling ontogenetic processes of teleost immunology. Finally, this study associated the first appearance of myeloid cells with an immune response on the molecular level: 2 days post fertilization embryos responded to LPS injection with upregulation of interleukin 1-beta, inducible nitric oxide synthase and serum amyloid A, and down-regulation of complement factor 3 and alpha2-macroglobulin, implying a functional embryonic innate defense system.
Novel immunoglobulin-like transcripts in teleost fish encode polymorphic receptors with cytoplasmic ITAM or ITIM an a new structural Ig domain similar to the natural cytotoxicity receptor NKp44
Stet, R.J.M. ; Hermsen, G.J. ; Westphal, A.H. ; Jukes, J. ; Engelsma, M.Y. ; Verburg-van Kemenade, B.M.L. ; Dortmans, J.C.F.M. ; Aveiro, J. ; Savelkoul, H.F.J. - \ 2005
Immunogenetics 57 (2005)1-2. - ISSN 0093-7711 - p. 77 - 89.
carp cyprinus-carpio - tumor-cell lysis - gene family - functional-characterization - histocompatibility complex - killer-cells - zebrafish - cluster - member - conservation
Members of the immunoglobulin superfamily (IgSF) include a group of innate immune receptors located in the leukocyte receptor complex (LRC) and other small clusters such as the TREM/NKp44 cluster. These receptors are characterised by the presence of immunoglobulin domains, a stalk, a transmembrane domain, and a cytoplasmic region containing either an immunoreceptor tyrosine-based inhibitory motif (ITIM) or are linked to an adapter molecule with an activation motif (ITAM) for downstream signalling. We have isolated two carp cDNA sequences encoding receptors in which the extracellular Ig domain structurally resembles the novel V-type Ig domain of NKp44. This is supported by a homology model. The cytoplasmic regions contain either an ITAM (Cyca-NILT1) or ITIMs (Cyca-NILT2). The tissue expression of these receptors is nearly identical, with the highest expression in the immunological organs. Peripheral blood leucocytes showed no detectable expression, but upon in vitro culture expressed NILT1, the activating receptor, and not the inhibitory NILT2 receptor. Southern blot analysis indicated that the NILT1 and NILT2 sequences belong to a multigene family. Analysis of the NILT Ig domain-encoding sequences amplified from both genomic DNA and cDNA revealed extensive haplotypic and allelic polymorphism. Database mining of the zebrafish genome identified several homologs on Chromosome 1, which also contains a cluster of class I major histocompatibility genes. This constellation is reminiscent of the TREM/NKp44 gene cluster and the HLA complex located on human Chromosome 6. The carp NILT genes form a unique cluster of innate immune receptors, which are highly polymorphic, and characterised by a new Ig structural subfamily and are distinct from the novel immune-type receptors (Nitrs) found in other fish species
Major histocompatibility genes in the Lake Tana African large barb species flock: evidence for complete partitioning of class II B, but nog class I, genes among different species
Kruiswijk, C.P. ; Hermsen, G.J. ; Heerwaarden, J. van; Dixon, B. ; Savelkoul, H.F.J. ; Stet, R.J.M. - \ 2005
Immunogenetics 56 (2005)12. - ISSN 0093-7711 - p. 894 - 908.
mhc class-i - nucleotide-sequences - oncorhynchus-mykiss - cichlid fishes - rainbow-trout - danio-rerio - polymorphism - zebrafish - evolution - locus
The 16 African large barb fish species of Lake Tana inhabit different ecological niches, exploit different food webs and have different temporal and spatial spawning patterns within the lake. This unique fish species flock is thought to be the result of adaptive radiation within the past 5 million years. Previous analyses of major histocompatibility class II B exon 2 sequences in four Lake Tana African large barb species revealed that these sequences are indeed under selection. No sharing of class II B alleles was observed among the four Lake Tana African large barb species. In this study we analysed the class II B exon 2 sequences of seven additional Lake Tana African large barb species and African large barbs from the Blue Nile and its tributaries. In addition, the presence and variability of major histocompatibility complex class I UA exon 3 sequences in six Lake Tana and Blue Nile African large barb species was analysed. Phylogenetic lineages are maintained by purifying or neutral selection on non-peptide binding regions. Class II B intron 1 and exon 2 sequences were not shared among the different Lake Tana African large barb species or with the riverine barb species. In contrast, identical class I UA exon 3 sequences were found both in the lacustrine and riverine barb species. Our analyses demonstrate complete partitioning of class II B alleles among Lake Tana African large barb species. In contrast, class I alleles remain for the large part shared among species. These different modes of evolution probably reflect the unlinked nature of major histocompatibility genes in teleost fishes.
Analysis of genomic and expressed major histocompatibility class Ia and class II genes in a hexaploid Lake Tana African 'large' barb individual (Barbus intermedius)
Kruiswijk, C.P. ; Hermsen, G.J. ; Fujiki, K. ; Dixon, B. ; Savelkoul, H.F.J. ; Stet, R.J.M. - \ 2004
Immunogenetics 55 (2004)11. - ISSN 0093-7711 - p. 770 - 781.
mhc class-i - complex class-i - carp cyprinus-carpio - beta-2-microglobulin gene - rainbow-trout - danio-rerio - beta(2)-microglobulin - fish - evolution - zebrafish
Expression of too many co-dominant major histocompatibility complex (MHC) alleles is thought to be detrimental to proper functioning of the immune system. Polyploidy of the genome will increase the number of expressed MHC genes unless they are prone to a silencing mechanism. In polyploid Xenopus species, the number of MHC class I and II genes has been physically reduced, as it does not increase with higher ploidy genomes. In the zebrafish some class IIB loci have been silenced, as only two genomically bona fide loci, DAA/DAB and DEA/DEB, have been described. Earlier studies indicated a reduction in the number of genomic and expressed class II MHC genes in a hexaploid African 'large' barb. This prompted us to study the number of MHC genes present in the genome of an African 'large' barb individual (Barbus intermedius) in relation to those expressed, adopting the following strategy. Full-length cDNA sequences were generated from mRNA and compared with partial genomic class Ia and II sequences generated by PCR using the same primer set. In addition, we performed Southern hybridizations to obtain a verification of the number of class I and IIB genes. Our study revealed three beta(2)-microglobulin, five class Ia, four class IIA, and four class IIB genes at the genomic level, which were shown to be expressed in the hexaploid barb individual. The class Ia and class II data indicate that the ploidy status does not correlate with the presence and expression of these MHC genes.