Expression profiling using random genomic DNA microarrays identifies differentially expressed genes associated with three major developmental stages of the protozoan parasite Leishmania major

Akopyants, Natalia S.; Matlib, Robin S.; Bukanova, Elena N.; Smeds, Matthew R.; Brownstein, Bernard H.; Stormo, Gary D.; Beverley, Stephen M.

doi:10.1016/j.molbiopara.2004.03.002

Cited by 108 publications

(85 citation statements)

References 54 publications

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“…This result agrees with that obtained by Akopyants et al who used Leishmania major genomic arrays similar to ours and looked for two-fold regulation [24]. When EST sequences were used to make the arrays and the boundary was set lower, the percentage of regulated genes appeared higher [28].…”

Section: Discussionsupporting

confidence: 92%

“…The original 927 total RNA experiment was less sensitive than the more recent one done using 427 total RNA, possibly because the more recent arrays are of superior quality. Akopyants et al [24], who compared mRNAs from three life-cycle stages of Leishmania using genomic microarrays, reported that the "efficiency" of recognition of a single regulated spot 9 (10) 9 (11) 5 (7) 23 (28) 427P 6 (9) 8 (11) 1 (7) 15 (27) Total spots 11 13 7 31 Details as in Table 2. For example, the experiment with 927 total RNA detected four ISG65 spots, four EP-PAG spots, and three MSP spots-a total of 11 for the three gene sets.…”

Section: Reliability and Reproducibilitymentioning

confidence: 99%

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The transcriptomes of Trypanosoma brucei Lister 427 and TREU927 bloodstream and procyclic trypomastigotes

Brems

Guilbride

Gundlesdodjir-Planck

et al. 2005

Molecular and Biochemical Parasitology

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We describe developmentally regulated genes in two strains of Trypanosoma brucei: the monomorphic strain Lister 427 and the pleomorphic strain TREU927. Expression patterns were obtained using an array of 24,567 genomic fragments. Probes were prepared from bloodstream-form or procyclic-form trypanosomes. Fourteen procyclic-specific and 77 bloodstream-specific signals were obtained from sequences matching variant surface glycoprotein or associated genes, and a further 17 regulated sequences were repetitive or transposable-element-related. Two hundred and eighty-six regulated spots corresponded to mRNAs from other protein-coding genes; these spots represent 191 different proteins. Regulation of 113 different genes (79 from procyclic forms, 34 from bloodstream-forms) was supported by at least two independent experiments or criteria; of these, about 60 were novel. Only two genes -encoding HSP83 and an importin-related protein -appeared to be regulated in the TREU927 strain only. Our results confirmed previous estimates that 2% of trypanosome genes show developmental regulation at the mRNA level.

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Section: Discussionsupporting

confidence: 92%

Section: Reliability and Reproducibilitymentioning

confidence: 99%

The transcriptomes of Trypanosoma brucei Lister 427 and TREU927 bloodstream and procyclic trypomastigotes

Brems

Guilbride

Gundlesdodjir-Planck

et al. 2005

Molecular and Biochemical Parasitology

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“…Early experience with small-scale arrays has demonstrated the potential value of this approach for studying gene expression in parasites [16][17][18][19]. We now report production and use of the first large-scale oligonucleotide microarray for a nematode parasite.…”

Section: Introductionmentioning

confidence: 95%

Profiling of gender-regulated gene transcripts in the filarial nematode Brugia malayi by cDNA oligonucleotide array analysis

Rush

Crosby

et al. 2005

Molecular and Biochemical Parasitology

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Microarray technology permits high-throughput comparisons of gene expression in different parasite stages or sexes and has been used widely. We report the first use of this technology for analysis of gene expression in filarial male and female worms. The slide array (comprised of 65-mer oligos representing 3569 EST clusters) was spotted with sequences selected from the extensive Brugia malayi EST database (http://zeldia.cap.ed.ac.uk/fgn/brugia.php). Arrays were hybridized with Cy dye labeled male and female cDNA. The experimental design included both biological and technical (dye-flip) replicates. The data were normalized for background and probe intensity, and the relative abundance of hybridized cDNA for each spot was determined. Genes showing two-fold or greater differences with P < 0.05 were considered gender-regulated candidates. One thousand one hundred and seventy of 2443 clusters (48%) with signals above threshold in at least one sex were considered as gender-regulated gene candidates. This included 520 and 650 clusters up-regulated in male and female worms, respectively. Fifty of 53 (94%) gender-regulated candidate genes identified by microarray analysis were confirmed by real-time RT-PCR. Approximately 61% of gender-regulated genes had significant similarity to known genes in other organisms such as Caenorhabditis elegans. Many C. elegans homologues of these genes have been reported to have reproductive phenotypes (sterility or abnormal embryo development) by RNA interference. This study has provided the first broad view of gender-regulated gene expression in B. malayi; this should lead to improved understanding of reproduction in filarial nematodes. More generally, this approach holds great promise as a means of studying stage-specific or tissue-specific gene expression in parasitic nematodes.

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“…El campo emergente de la genómica funcional está dedicado al desarrollo y la aplicación de métodos que permitan estudiar de manera eficiente la expresión de varios genes simultáneamente, por las tecnologías de microarreglos y proteómica (119)(120)(121).…”

Section: Conclusiones Y Perspectivasunclassified

Manipulación genética y el estudio del parásito protozoario Leishmania.

Cortázar

Walker

2004

biomedica

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Durante los últimos 15 años se ha dado paso al entendimiento de muchos aspectos de la genómica funcional de Leishmania gracias a los avances en la metodología de transfección de ADN dentro de la célula de este protozoario, la eliminación y la complementación de genes por medio de recombinación homóloga y las estrategias para la selección de células transfectadas. Estos acercamientos tienen el potencial de brindar información sobre la expresión génica y la función de las proteínas en el contexto del parásito intacto. Dado que el genoma de Leishmania muestra una carencia acentuada de los factores conocidos de iniciación de la transcripción y que la expresión génica está regulada casi completamente a nivel postranscripcional (a través del empalme de los ARNm y los mecanismos que involucran el procesamiento diferencial de la región no traducida 3' del ARNm (3'UTR), la transfección génica representa una herramienta útil para la identificación y el análisis funcional de los genes de interés así como de los mecanismos que dirigen su regulación. El desarrollo de los sistemas de manipulación genética también ha abierto nuevos horizontes para la identificación de genes esenciales involucrados en la virulencia, la supervivencia intracelular y la resistencia a drogas de Leishmania, así como para la validación de proteínas específicas del parásito como nuevos blancos quimio e inmunoterapéuticos. En esta revisión presentamos los avances más recientes en el campo de la manipulación genética en Leishmania, los cuales permiten análisis estructurales, funcionales y de fenotipo, por medio de la eliminación y complementación génica a través de la transfección transitoria o permanente de genes en este parásito.Palabras clave: Leishmania, transfección de ADN, expresión génica, eliminación y complementación génica, genómica funcional. Genetic manipulation and the study of the protozoan parasite LeishmaniaDuring the last 15 years, many aspects of the functional genomics of Leishmania have been revealed due to advances in DNA transfection, gene disruption and complementation through homologous recombination, and efficient strategies for the selection of transfected cells. These strategies have provided information about gene expression and protein function in the context of the intact parasite. The genome of Leishmania shows a marked deficiency of known transcription initiation factors, and gene expression is regulated almost entirely at the posttranscriptional level through trans-splicing of mRNAs and novel control mechanisms involving differential processing of 3' -untranslated regions (3'-UTRs) of mRNAs. Therefore, gene transfection represents a useful tool for the identification and functional analysis of genes of interest as well as the mechanisms that direct their regulation. The development of genetic manipulation systems has provided opportunities for the study of genes involved in virulence, intracellular survival and drug resistance of Leishmania, as well as for the functional validation of specific parasite proteins as new ch...

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Expression profiling using random genomic DNA microarrays identifies differentially expressed genes associated with three major developmental stages of the protozoan parasite Leishmania major

Cited by 108 publications

References 54 publications

The transcriptomes of Trypanosoma brucei Lister 427 and TREU927 bloodstream and procyclic trypomastigotes

The transcriptomes of Trypanosoma brucei Lister 427 and TREU927 bloodstream and procyclic trypomastigotes

Profiling of gender-regulated gene transcripts in the filarial nematode Brugia malayi by cDNA oligonucleotide array analysis

Manipulación genética y el estudio del parásito protozoario Leishmania.

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