Gene expression analysis of an integrin family of genes by systematic multiplex reverse transcription‐polymerase chain reaction

Yamamoto, Miyako; Yamamoto, Ami; Leung, Patricia C.; Yamamoto, Fumiichiro

doi:10.1002/elps.200305952

Cited by 6 publications

(4 citation statements)

References 26 publications

(27 reference statements)

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“…We prepared cDNA by reverse-transcription using oligo dT primers and the Advantage RT-for-PCR Kit (BD Biosciences-Clontech). We followed the SM RT-PCR experimental protocols described previously [29], [50], [51], [52]. Complementary DNA samples were used as templates to examine gene expression.…”

Section: Methodsmentioning

confidence: 99%

Rare and Frequent Promoter Methylation, Respectively, of TSHZ2 and 3 Genes That Are Both Downregulated in Expression in Breast and Prostate Cancers

et al. 2011

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BackgroundNeoplastic cells harbor both hypomethylated and hypermethylated regions of DNA. Whereas hypomethylation is found mainly in repeat sequences, regional hypermethylation has been linked to the transcriptional silencing of certain tumor suppressor genes. We attempted to search for candidate genes involved in breast/prostate carcinogenesis, using the criteria that they should be expressed in primary cultures of normal breast/prostate epithelial cells but are frequently downregulated in breast/prostate cancer cell lines and that their promoters are hypermethylated.Methodology/Principal FindingsWe identified several dozens of candidates among 194 homeobox and related genes using Systematic Multiplex RT-PCR and among 23,000 known genes and 23,000 other expressed sequences in the human genome by DNA microarray hybridization. An additional examination, by real-time qRT-PCR of clinical specimens of breast cancer, further narrowed the list of the candidates. Among them, the most frequently downregulated genes in tumors were NP_775756 and ZNF537, from the homeobox gene search and the genome-wide search, respectively. To our surprise, we later discovered that these genes belong to the same gene family, the 3-member Teashirt family, bearing the new names of TSHZ2 and TSHZ3. We subsequently determined the methylation status of their gene promoters. The TSHZ3 gene promoter was found to be methylated in all the breast/prostate cancer cell lines and some of the breast cancer clinical specimens analyzed. The TSHZ2 gene promoter, on the other hand, was unmethylated except for the MDA-MB-231 breast cancer cell line. The TSHZ1 gene was always expressed, and its promoter was unmethylated in all cases.Conclusions/SignificanceTSHZ2 and TSHZ3 genes turned out to be the most interesting candidates for novel tumor suppressor genes. Expression of both genes is downregulated. However, differential promoter methylation suggests the existence of distinctive mechanisms of transcriptional inactivation for these genes.

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Section: Methodsmentioning

confidence: 99%

Rare and Frequent Promoter Methylation, Respectively, of TSHZ2 and 3 Genes That Are Both Downregulated in Expression in Breast and Prostate Cancers

et al. 2011

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“…We then developed the SM RT-PCR systems of 39 human Hox genes [14] and 26 human integrin subunit genes [15]. Using these systems, we have demonstrated that SM RT-PCR is a useful method of semiquantitative quantification.…”

Section: Introductionmentioning

confidence: 99%

“…During the past several years, we have been developing a novel technique for semiquantitative quantification of gene expression named systematic multiplex reverse transcription-PCR (SM RT-PCR) [13][14][15]. SM RT-PCR is distinguishable from standard multiplex RT-PCR because it utilizes primers that amplify sequences that fall within a single exon of genes.…”

Section: Introductionmentioning

confidence: 99%

Scanning copy number and gene expression on the 16p13.3‐13.2 chromosomal region by the systematic multiplex polymerase chain reaction and reverse transcription‐polymerase chain reaction methods

2006

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We developed the systematic multiplex reverse transcription-PCR (SM RT-PCR) method that is distinguishable from other multiplex RT-PCR methods by optimized PCR conditions allowing amplification of sequences that fall within a single exon of genes of similar band intensity using genomic DNA template as a calibration standard. Using an SM RT-PCR system of proto-oncogenes and tumor suppressor genes, we previously showed that the SM RT-PCR system, which was developed for cDNA expression analysis, could also be used for a more exquisite analysis of copy number changes in genomic DNA. Here we report that the SM PCR method semiquantitatively detected less than a two-fold difference in copy number. Furthermore, we also report the results of subchromosomal scanning of copy number and expression using the SM PCR and SM RT-PCR methods. We identified and characterized the novel homozygous deletion that spans over 12-plus genes on 16p13.3-13.2 in the MDA-MB-468 breast cancer cell line.

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“…In mammals, 18 α and 8 β subunits combine to form 24 integrins [ 1 ]. Alternative splicing [ 2 ] and post-translational modifications [ 3 ] ensure a high diversity in integrin structure. Such diversity in subunit composition contributes to their expanded ligand recognition, binding to the extracellular matrix (ECM), and coupling to downstream signaling pathways [ 4 , 5 , 6 , 7 , 8 , 9 ].…”

Section: Introductionmentioning

confidence: 99%

Novel Functions of Integrins as Receptors of CD154: Their Role in Inflammation and Apoptosis

Hassan

Salti

Mourad

2022

Cells

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CD154, an inflammatory mediator also known as CD40 ligand, has been identified as a novel binding partner for some members of the integrin family. The αIIbβ3, specifically expressed on platelets, was the first integrin to be described as a receptor for CD154 after CD40. Its interaction with soluble CD154 (sCD154) highly contributes to thrombus formation and stability. Identifying αIIbβ3 opened the door for investigating other integrins as partners of CD154. The αMβ2 expressed on myeloid cells was shown capable of binding CD154 and contributing as such to cell activation, adhesion, and release of proinflammatory mediators. In parallel, α5β1 communicates with sCD154, inducing pro-inflammatory responses. Additional pathogenic effects involving apoptosis-preventing functions were exhibited by the CD154–α5β1 dyad in T cells, conferring a role for such interaction in the survival of malignant cells, as well as the persistence of autoreactive T cells. More recently, CD154 receptors integrated two new integrin members, αvβ3 and α4β1, with little known as to their biological significance in this context. This article provides an overview of the novel role of integrins as receptors of CD154 and as critical players in pro-inflammatory and apoptotic responses.

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Gene expression analysis of an integrin family of genes by systematic multiplex reverse transcription‐polymerase chain reaction

Cited by 6 publications

References 26 publications

Rare and Frequent Promoter Methylation, Respectively, of TSHZ2 and 3 Genes That Are Both Downregulated in Expression in Breast and Prostate Cancers

Rare and Frequent Promoter Methylation, Respectively, of TSHZ2 and 3 Genes That Are Both Downregulated in Expression in Breast and Prostate Cancers

Scanning copy number and gene expression on the 16p13.3‐13.2 chromosomal region by the systematic multiplex polymerase chain reaction and reverse transcription‐polymerase chain reaction methods

Novel Functions of Integrins as Receptors of CD154: Their Role in Inflammation and Apoptosis

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