Biomedical subjects

Fredrik Sterky

Publications and source records attributed to Fredrik Sterky.

13 recordsLinked to original sources

PubMed2005

A human protein atlas for normal and cancer tissues based on antibody proteomics.

Antibody-based proteomics provides a powerful approach for the functional study of the human proteome involving the systematic generation of protein-specific affinity reagents. We used this strategy to construct a comprehensive, antibody-based protein atlas for expression and localization profiles in 48 normal human tissues and 20 different cancers. Here we report a new publicly available database containing, in the first version, approximately 400,000 high resolution images corresponding to more than 700 antibodies toward human proteins. Each image has been annotated by a certified pathologist to provide a knowledge base for functional studies and to allow queries about protein profiles in normal and disease tissues. Our results suggest it should be possible to extend this analysis to the majority of all human proteins thus providing a valuable tool for medical and biological research.

Antibodies↗

PubMed2005

The genome sequence of black cottonwood (Populus trichocarpa) reveals 18 conserved cellulose synthase (CesA) genes.

The genome sequence of Populus trichocarpa was screened for genes encoding cellulose synthases by using full-length cDNA sequences and ESTs previously identified in the tissue specific cDNA libraries of other poplars. The data obtained revealed 18 distinct CesA gene sequences in P. trichocarpa. The identified genes were grouped in seven gene pairs, one group of three sequences and one single gene. Evidence from gene expression studies of hybrid aspen suggests that both copies of at least one pair, CesA3-1 and CesA3-2, are actively transcribed. No sequences corresponding to the gene pair, CesA6-1 and CesA6-2, were found in Arabidopsis or hybrid aspen, while one homologous gene has been identified in the rice genome and an active transcript in Populus tremuloides. A phylogenetic analysis suggests that the CesA genes previously associated with secondary cell wall synthesis originate from a single ancestor gene and group in three distinct subgroups. The newly identified copies of CesA genes in P. trichocarpa give rise to a number of new questions concerning the mechanism of cellulose synthesis in trees.

Chromosome Mapping↗

PubMed2005

SNP discovery using advanced algorithms and neural networks.

UNLABELLED: Forage is an application which uses two neural networks for detecting single nucleotide polymorphisms (SNPs). Potential SNP candidates are identified in multiple alignments. Each candidate is then represented by a vector of features, which is classified as SNP or monomorphic by the networks. A validated dataset of SNPs was constructed from experimentally verified SNP data and used for network training and method evalutation. AVAILABILITY: The package is available at biobase.biotech.kth.se/forage/

Algorithms↗

PubMed2005

Carbohydrate-active enzymes involved in the secondary cell wall biogenesis in hybrid aspen.

Wood formation is a fundamental biological process with significant economic interest. While lignin biosynthesis is currently relatively well understood, the pathways leading to the synthesis of the key structural carbohydrates in wood fibers remain obscure. We have used a functional genomics approach to identify enzymes involved in carbohydrate biosynthesis and remodeling during xylem development in the hybrid aspen Populus tremula x tremuloides. Microarrays containing cDNA clones from different tissue-specific libraries were hybridized with probes obtained from narrow tissue sections prepared by cryosectioning of the developing xylem. Bioinformatic analyses using the sensitive tools developed for carbohydrate-active enzymes allowed the identification of 25 xylem-specific glycosyltransferases belonging to the Carbohydrate-Active EnZYme families GT2, GT8, GT14, GT31, GT43, GT47, and GT61 and nine glycosidases (or transglycosidases) belonging to the Carbohydrate-Active EnZYme families GH9, GH10, GH16, GH17, GH19, GH28, GH35, and GH51. While no genes encoding either polysaccharide lyases or carbohydrate esterases were found among the secondary wall-specific genes, one putative O-acetyltransferase was identified. These wood-specific enzyme genes constitute a valuable resource for future development of engineered fibers with improved performance in different applications.

Cell Wall↗

PubMed2005

EST data suggest that poplar is an ancient polyploid.

We analysed the publicly available expressed sequence tag (EST) collections for the genus Populus to examine whether evidence can be found for large-scale gene-duplication events in the evolutionary past of this genus. The ESTs were clustered into unigenes for each poplar species examined. Gene families were constructed for all proteins deduced from these unigenes, and K(S) dating was performed on all paralogs within a gene family. The fraction of paralogs was then plotted against the K(S) values, which resulted in a distribution reflecting the age of duplicated genes in poplar. Sufficient EST data were available for seven different poplar species spanning four of the six sections of the genus Populus. For all these species, there was evidence that a large-scale gene-duplication event had occurred. From our analysis it is clear that all poplar species have shared the same large-scale gene-duplication event, suggesting that this event must have occurred in the ancestor of poplar, or at least very early in the evolution of the Populus genus.

Biological Evolution↗

PubMed2005

Selection of protein epitopes for antibody production.

Protein functional analysis in the post-genomic era is a huge task that has to be approached by different methods in parallel. The use of protein-specific antibodies in conjunction with tissue microarrays has proven to be one important technology. In this study, we present a strategy for the optimized design of protein subfragments for subsequent antibody production. The fragments are selected based on a principle of lowest sequence similarity to other human proteins, optimally to generate antibodies with high selectivity. Furthermore, the fragments should have properties optimized for efficient protein production in Escherichia coli. The strategy has been implemented in Bishop, which is a Java-based software enabling the high-throughput production of protein fragments. Bishop allows for the avoidance of certain restriction enzyme sites, transmembrane regions, and signal peptides. A Basic Local Alignment Search Tool (BLAST) scanning procedure permits the selection of fragments of a selected size with a minimal sequence similarity to other proteins. The software and the strategy were evaluated on a human test data set and verified to fulfill the requested criteria.

Algorithms↗

PubMed2004

A Populus EST resource for plant functional genomics.

Trees present a life form of paramount importance for terrestrial ecosystems and human societies because of their ecological structure and physiological function and provision of energy and industrial materials. The genus Populus is the internationally accepted model for molecular tree biology. We have analyzed 102,019 Populus ESTs that clustered into 11,885 clusters and 12,759 singletons. We also provide >4,000 assembled full clone sequences to serve as a basis for the upcoming annotation of the Populus genome sequence. A public web-based EST database (POPULUSDB) provides digital expression profiles for 18 tissues that comprise the majority of differentiated organs. The coding content of Populus and Arabidopsis genomes shows very high similarity, indicating that differences between these annual and perennial angiosperm life forms result primarily from differences in gene regulation. The high similarity between Populus and Arabidopsis will allow studies of Populus to directly benefit from the detailed functional genomic information generated for Arabidopsis, enabling detailed insights into tree development and adaptation. These data will also valuable for functional genomic efforts in Arabidopsis.

Ecosystem↗

PubMed2004

Expansins abundant in secondary xylem belong to subgroup A of the alpha-expansin gene family.

Differentiation of xylem cells in dicotyledonous plants involves expansion of the radial primary cell walls and intrusive tip growth of cambial derivative cells prior to the deposition of a thick secondary wall essential for xylem function. Expansins are cell wall-residing proteins that have an ability to plasticize the cellulose-hemicellulose network of primary walls. We found expansin activity in proteins extracted from the cambial region of mature stems in a model tree species hybrid aspen (Populus tremula x Populus tremuloides Michx). We identified three alpha-expansin genes (PttEXP1, PttEXP2, and PttEXP8) and one beta-expansin gene (PttEXPB1) in a cambial region expressed sequence tag library, among which PttEXP1 was most abundantly represented. Northern-blot analyses in aspen vegetative organs and tissues showed that PttEXP1 was specifically expressed in mature stems exhibiting secondary growth, where it was present in the cambium and in the radial expansion zone. By contrast, PttEXP2 was mostly expressed in developing leaves. In situ reverse transcription-PCR provided evidence for accumulation of mRNA of PttEXP1 along with ribosomal rRNA at the tips of intrusively growing xylem fibers, suggesting that PttEXP1 protein has a role in intrusive tip growth. An examination of tension wood and leaf cDNA libraries identified another expansin, PttEXP5, very similar to PttEXP1, as the major expansin in developing tension wood, while PttEXP3 was the major expansin expressed in developing leaves. Comparative analysis of expansins expressed in woody stems in aspen, Arabidopsis, and pine showed that the most abundantly expressed expansins share sequence similarities, belonging to the subfamily A of alpha-expansins and having two conserved motifs at the beginning and end of the mature protein, RIPVG and KNFRV, respectively. This conservation suggests that these genes may share a specialized, not yet identified function.

Amino Acid Sequence↗

PubMed2004

A transcriptional timetable of autumn senescence.

BACKGROUND: We have developed genomic tools to allow the genus Populus (aspens and cottonwoods) to be exploited as a full-featured model for investigating fundamental aspects of tree biology. We have undertaken large-scale expressed sequence tag (EST) sequencing programs and created Populus microarrays with significant gene coverage. One of the important aspects of plant biology that cannot be studied in annual plants is the gene activity involved in the induction of autumn leaf senescence. RESULTS: On the basis of 36,354 Populus ESTs, obtained from seven cDNA libraries, we have created a DNA microarray consisting of 13,490 clones, spotted in duplicate. Of these clones, 12,376 (92%) were confirmed by resequencing and all sequences were annotated and functionally classified. Here we have used the microarray to study transcript abundance in leaves of a free-growing aspen tree (Populus tremula) in northern Sweden during natural autumn senescence. Of the 13,490 spotted clones, 3,792 represented genes with significant expression in all leaf samples from the seven studied dates. CONCLUSIONS: We observed a major shift in gene expression, coinciding with massive chlorophyll degradation, that reflected a shift from photosynthetic competence to energy generation by mitochondrial respiration, oxidation of fatty acids and nutrient mobilization. Autumn senescence had much in common with senescence in annual plants; for example many proteases were induced. We also found evidence for increased transcriptional activity before the appearance of visible signs of senescence, presumably preparing the leaf for degradation of its components.

Aging↗

PubMed2004

MYB transcription factors are differentially expressed and regulated during secondary vascular tissue development in hybrid aspen.

More than 120,000 poplar ESTs have been sequenced from 20 different cDNA libraries by the Swedish Centre for Tree Functional Genomics. We screened this EST collection for MYB transcription factors involved in secondary vascular tissue formation, and genes assigned as PttMYB3Ra, PttMYB4a and PttMYB21a were selected for further characterisation. Three MYB genes showed different expression patterns in various organs, tissues and stem sub-sections representing different developmental stages of vascular tissue formation. Furthermore, the analysis showed that PttMYB21a expression was much higher in secondary cell wall formation zone of xylem and phloem fibers than in other developmental zones. Transgenic hybrid aspen plants, expressing the 3'-part of the PttMYB21a gene in antisense orientation were generated to assess the function of PttMYB21a gene in vascular tissue formation and lignification. All transgenic lines showed reduced growth and had fewer internodes compared to the wild-type. The analysis of selected lines showed that acid soluble lignin present in the bark was higher in transgenic lines as compared to wild-type plants. Moreover a higher transcript level of caffeoyl-CoA 3-O-methyltransferase [CCoAOMT]; EC 2.1.1.104) was found in the phloem of the transgenic plants, suggesting that PttMYB21a might function as a transcriptional repressor.

Amino Acid Sequence↗

PubMed2003

Gene expression in autumn leaves.

Two cDNA libraries were prepared, one from leaves of a field-grown aspen (Populus tremula) tree, harvested just before any visible sign of leaf senescence in the autumn, and one from young but fully expanded leaves of greenhouse-grown aspen (Populus tremula x tremuloides). Expressed sequence tags (ESTs; 5,128 and 4,841, respectively) were obtained from the two libraries. A semiautomatic method of annotation and functional classification of the ESTs, according to a modified Munich Institute of Protein Sequences classification scheme, was developed, utilizing information from three different databases. The patterns of gene expression in the two libraries were strikingly different. In the autumn leaf library, ESTs encoding metallothionein, early light-inducible proteins, and cysteine proteases were most abundant. Clones encoding other proteases and proteins involved in respiration and breakdown of lipids and pigments, as well as stress-related genes, were also well represented. We identified homologs to many known senescence-associated genes, as well as seven different genes encoding cysteine proteases, two encoding aspartic proteases, five encoding metallothioneins, and 35 additional genes that were up-regulated in autumn leaves. We also indirectly estimated the rate of plastid protein synthesis in the autumn leaves to be less that 10% of that in young leaves.

Arabidopsis Proteins↗

PubMed2002

Molecular cloning and characterization of a cDNA encoding poplar UDP-glucose dehydrogenase, a key gene of hemicellulose/pectin formation.

Plant UDP-glucose dehydrogenase (UGDH) is an important enzyme in the formation of hemicellulose and pectin, the components of newly formed cell walls. A cDNA clone (Ugdh) corresponding to UGDH was isolated from a cDNA library prepared from cambial zone of poplar (Populus tremula x tremuloides). Within the 1824-nucleotide (nt)-long clone, an open reading frame encoded a protein of 481 amino acids (aa), with a calculated molecular weight of 53.1 kDa. The derived aa sequence showed 90% and 63% identity with UGDHs from soybean and bovine liver, respectively, and had highly conserved aa motifs believed to be of importance for nt binding and catalytic efficiency. In poplar, the Ugdh corresponds to one or two genes, as found by genomic Southern analysis. The gene was expressed predominantly in differentiating xylem and young leaves, with little expression in the phloem zone of the stem. The expression pattern matched that of UGDH protein, as found by immunoblotting. In leaves, the Ugdh expression was upregulated by a short-term feeding with sucrose, sorbitol and polyethylene glycol, and this effect was to some extent mimicked by light exposure. The data suggest that Ugdh is regulated via an osmoticum-dependent pathway, possibly related to the availability of osmotically active carbohydrate precursors to UDP-glucose, a substrate of UGDH.

Amino Acid Sequence↗

PubMed2002

Microarray data quality analysis: lessons from the AFGC project. Arabidopsis Functional Genomics Consortium.

Genome-wide expression profiling with DNA microarrays has and will provide a great deal of data to the plant scientific community. However, reliability concerns have required the development data quality tests for common systematic biases. Fortunately, most large-scale systematic biases are detectable and some are correctable by normalization. Technical replication experiments and statistical surveys indicate that these biases vary widely in severity and appearance. As a result, no single normalization or correction method currently available is able to address all the issues. However, careful sequence selection, array design, experimental design and experimental annotation can substantially improve the quality and biological of microarray data. In this review, we discuss these issues with reference to examples from the Arabidopsis Functional Genomics Consortium (AFGC) microarray project.

Arabidopsis↗