The monogeneans are external parasites mostly of fish with life cycles consisting of a free-swimming larva that attaches to a fish to begin transformation to the parasitic adult form. They have only one host during their life, typically of just one species. The worms may produce enzymes that digest the host tissues or graze on surface mucus and skin particles.
Most monogeneans are hermaphroditic, but the sperm develop first, and it is typical for them to mate between individuals and not to self-fertilize. The trematodes, or flukes, are internal parasites of mollusks and many other groups, including humans. Trematodes have complex life cycles that involve a primary host in which sexual reproduction occurs and one or more secondary hosts in which asexual reproduction occurs.
The primary host is almost always a mollusk. Trematodes are responsible for serious human diseases including schistosomiasis, caused by a blood fluke Schistosoma. The disease infects an estimated million people in the tropics and leads to organ damage and chronic symptoms including fatigue. Infection occurs when a human enters the water, and a larva, released from the primary snail host, locates and penetrates the skin.
The parasite infects various organs in the body and feeds on red blood cells before reproducing. Many of the eggs are released in feces and find their way into a waterway where they are able to reinfect the primary snail host. The cestodes, or tapeworms, are also internal parasites, mainly of vertebrates. Tapeworms live in the intestinal tract of the primary host and remain fixed using a sucker on the anterior end, or scolex, of the tapeworm body.
The remaining body of the tapeworm is made up of a long series of units called proglottids, each of which may contain an excretory system with flame cells, but will contain reproductive structures, both male and female.
Most reproduction occurs by cross-fertilization. The proglottid detaches and is released in the feces of the host. The fertilized eggs are eaten by an intermediate host. The juvenile worms emerge and infect the intermediate host, taking up residence, usually in muscle tissue.
When the muscle tissue is eaten by the primary host, the cycle is completed. There are several tapeworm parasites of humans that are acquired by eating uncooked or poorly cooked pork, beef, and fish. The phylum Nematoda , or roundworms, includes more than 28, species with an estimated 16, parasitic species. Most nematodes look similar to each other: slender tubes, tapered at each end [Figure 3].
Nematodes are pseudocoelomates and have a complete digestive system with a distinct mouth and anus. The nematode body is encased in a cuticle, a flexible but tough exoskeleton, or external skeleton, which offers protection and support. The cuticle contains a carbohydrate-protein polymer called chitin. The cuticle also lines the pharynx and rectum.
Although the exoskeleton provides protection, it restricts growth, and therefore must be continually shed and replaced as the animal increases in size. There may also be a sharp stylet that can protrude from the mouth to stab prey or pierce plant or animal cells. The mouth leads to a muscular pharynx and intestine, leading to the rectum and anal opening at the posterior end.
In nematodes, the excretory system is not specialized. Nitrogenous wastes are removed by diffusion. In marine nematodes, regulation of water and salt is achieved by specialized glands that remove unwanted ions while maintaining internal body fluid concentrations. Most nematodes have four nerve cords that run along the length of the body on the top, bottom, and sides.
Beneath the epidermis lies a layer of longitudinal muscles that permits only side-to-side, wave-like undulation of the body. View this video to see nematodes move about and feed on bacteria. Nematodes employ a diversity of sexual reproductive strategies depending on the species; they may be monoecious, dioecious separate sexes , or may reproduce asexually by parthenogenesis.
Caenorhabditis elegans is nearly unique among animals in having both self-fertilizing hermaphrodites and a male sex that can mate with the hermaphrodite. Arthropoda dominate the animal kingdom with an estimated 85 percent of known species, with many still undiscovered or undescribed. The principal characteristics of all the animals in this phylum are functional segmentation of the body and the presence of jointed appendages [Figure 4].
As members of Ecdysozoa, arthropods also have an exoskeleton made principally of chitin. Arthropoda is the largest phylum in the animal world in terms of numbers of species, and insects form the single largest group within this phylum. Arthropods are true coelomate animals and exhibit prostostomic development. A unique feature of arthropods is the presence of a segmented body with fusion of certain sets of segments to give rise to functional segments. Fused segments may form a head, thorax, and abdomen, or a cephalothorax and abdomen, or a head and trunk.
The coelom takes the form of a hemocoel or blood cavity. The open circulatory system, in which blood bathes the internal organs rather than circulating in vessels, is regulated by a two-chambered heart. The worm sorter records a fluorescence profile of each worm in the form of a curve, which reflects the intensity of GFP from the mouth to the tail. The mean of all values for each sample was determined, and the average of each duplicate was calculated.
At this point, worms were transferred to fresh RNAi plates containing 1. Worms developed to the gravid stage and laid eggs. After 24 h of egg-laying, the P 0 worms all in the gravid stage were discarded in order to prevent additional eggs from being laid. On day 5, both the total number of surviving larvae and the number of unhatched eggs were counted. ZnMP fluorescence intensity was measured as described previously [12]. Worms were washed from bombardment plates and transferred to plates seeded with a lawn of E.
Individual transgenic lines were isolated and transferred to axenic liquid mCeHR-2 medium supplemented with antibiotics. After two weeks of serial passages, worms were bleached and maintained as transgenic strains in axenic liquid mCeHR-2 medium. The microarray data was submitted to GEO on Aug 6, A compilation of heat maps generated following normalization of the data see Figure 2 legend using GeneSpring v7.
Yellow represents no change in signal intensity, blue indicates a decrease, and red indicates an increase in signal intensity. Of the hrg s identified in the study, were annotated with a biological process. Genes were analyzed using the Fisher's exact test and the topGO package from R. The most significant GO terms and their associated parent terms were used to construct a hierarchical graph such that the specificity of the terms increased as we moved from top to bottom.
The complete table of P- values and a full description of the GO term associated with each gene can be found in Tables S7 , S8 , and S9. Data was collected using the Affymetrix C. Each entry in the table represents a gene whose expression changed at least 1.
The table has six columns for each hrg. If the column is blank, then the change was less than 1. Heme-responsive genes whose expression is upregulated greater than 1.
Heme-responsive genes used to corroborate the microarray results. Of the hrgs whose expression changed significantly in response to heme, the results of a gene ontology analysis were used to assign a known biological process and molecular function to 63 genes. Each GO ID is assigned a unique function or association. Green shading indicates that term was included in the corresponding GO enrichment figures. The algorithms available on the Kyoto Encyclopedia of Genes and Genomes website were used to make functional predictions for each of the hrg s identified in the microarray.
Ten hrg s were mapped to KEGG pathways. Phenotypes observed when hrg s were knocked down in experiments performed by other laboratories and compiled on Wormbase. Negative fold change implies down regulation. Worm strains were provided by the Caenorhabditis Genetics Center. Abstract Heme is a cofactor in proteins that function in almost all sub-cellular compartments and in many diverse biological processes.
Author Summary Heme is an iron-containing cofactor for proteins involved in many critical cellular processes. Chisholm, University of California San Diego, United States of America Received: January 14, ; Accepted: June 30, ; Published: July 29, This is an open-access article distributed under the terms of the Creative Commons Public Domain declaration which stipulates that, once placed in the public domain, this work may be freely reproduced, distributed, transmitted, modified, built upon, or otherwise used by anyone for any lawful purpose.
Introduction From a nutritional perspective, heme is a readily bioavailable source of iron for human consumption [1] , [2]. Results Strategy to profile genes that are transcriptionally regulated by heme in C. Download: PPT. Figure 1. Profiling strategy for analysis of heme-responsive genes. Identification of hrg s in C. Comparative genome analyses of hrg s in vertebrates and parasites Since identification of the hrg s common to both C. Figure 5. Depiction of the number of hrgs found on each chromosome relative to the number of megabases in that chromosome.
Figure 6. Chromosomal location and fold-change of each heme-responsive gene. Functional validation reveals novel hrg s that are essential for heme homeostasis in C.
Figure 8. Characterization of the three candidate genes identified from the functional RNAi screens. Discussion A major impediment to the identification of heme uptake and transport pathways has been the inability to disassociate the tightly regulated process of heme synthesis from the downstream pathways for heme transport [3].
Materials and Methods Biological materials, strains, and worm culture C. Microarray data analysis Expression data were normalized and analyzed using MAS 5. Generation of the hrg mini-library The Ahringer and Vidal feeding libraries were replicated to individual well plates [50] , [51]. Supporting Information. Figure S1. Heat map for the heme microarrays. Figure S2. Gene ontology GO enrichment analysis of heme-responsive genes. Table S1. The heme-responsive genes identified by the microarray.
Table S2. Table S3. Table S4. Table S5. Heme-responsive genes with known Gene Ontology terms. Table S6. Table S7. Table S8. Table S9. Table S Heme-responsive genes assigned to a biological pathway by KEGG analysis. Previously reported RNAi phenotypes of heme-responsive genes. Heme-responsive genes with predicted TMDs. References 1. Semin Hematol 27— View Article Google Scholar 2. World J Gastroenterol — View Article Google Scholar 3. Chem Rev. Hamza I Intracellular trafficking of porphyrins.
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Nat Genet 25— Bioinformatics — In Silico Biol 5: — Thomas JH Analysis of homologous gene clusters in Caenorhabditis elegans reveals striking regional cluster domains. Genetics — J Proteome Res. Proteins — Furthermore, the conclusion in Liu et al. In the present study, we have provided strong experimental evidence that these soybean pre-miR are functional in soybean roots. Also, MIR gene family members exhibited varying degrees of expression in young soybean roots Supplementary Fig.
S2A, B , which is consistent with previous findings Li et al. However, Li et al. Tian et al. We found that all canonical pre-miR and miR are down-regulated early during the syncytium formation phase, and up-regulated during the maintenance phase Fig. Thus, our finding that expression patterns differ drastically at different time points i. S2A, C. Moreover, Li et al. Eleven soybean GRF genes are specifically up-regulated during the cumulative syncytium formation phase at 8 dpi , which in our infection system was between 5 and 13 dpi Fig.
Moreover, promoter analyses Fig. Microarray analysis was previously performed on laser capture-microdissected H. However, this study did not present any data on GRF genes, which is probably due to a number of factors.
For instance, the only genes that were analyzed in the latter study were those that first changed in expression at 2 dpi, and then those genes were subsequently analyzed at 5 and 10 dpi. Thus, it is likely that at 2 dpi GRF genes are not yet up-regulated to the point of detection, which would be consistent with our results Fig. Other microarray analyses were also performed on H. Lack of data presented for GRF genes in these studies could have also been due to insufficient representation on the GeneChip Ithal et al.
Also, many other microarray and RNA-seq studies have been performed on H. In the RAM, stem cell progeny undergo rapid cell division to ensure that there are enough cells for proper growth, and these rapidly dividing cells are called the transit-amplifying cells TACs Rodriguez et al. Thus, the function of the miR— GRF regulatory network in soybean roots appears to be similar to that of other plant species Bazin et al.
Conversely, during the syncytium maintenance phase, GRF genes are post-transcriptionally down-regulated by the de-repressed miR expression Figs 1B , 3E—H , 4. Hence, soybean GRF genes appear to be regulated in the H. S4 , has no promoter activity in H. Thus, the developmental program of developing H. When plants are under high pathogen stress, resources are devoted towards defense responses, while growth is stunted and development is delayed.
This phenomenon is known as the growth—defense trade-off Huot et al. GRFs have been implicated in various abiotic and biotic stress conditions Liu et al. GRFs are thus hypothesized to co-ordinate the interactions between defense signaling and growth and developmental pathways Liu et al.
In this context, GRFs could be thought to promote growth by maintaining rapid cell cycles while simultaneously suppressing defense responses. This phenotype is probably reflected by a decreased elongation zone Rodriguez et al. Thus, although GRF genes are required to maintain proper soybean lateral root numbers, and productive H.
Feedback regulation of miRNAs by their transcription factor targets has been demonstrated in several studies Gutierrez et al. This complex feedback loop ensures a precise transcriptional equilibrium.
In summary, we have investigated if a miR— GRF regulatory system operates in the agronomically important interaction between H. Thus, by specifically interfering with this network in syncytia using an H. Table S2. This work was supported by grants from the Iowa Soybean Association. The authors would like to thank Tom Maier and Danielle Sill for technical assistance. The authors declare no conflict of interest. Timecourse microarray analyses reveal global changes in gene expression of susceptible Glycine max soybean roots during infection by Heterodera glycines soybean cyst nematode.
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