PrimerSeq: Design and Visualization of RT-PCR Primers for Alternative Splicing Using RNA-seq Data

The vast majority of multi-exon genes in higher eukaryotes are alternatively spliced and changes in alternative splicing（AS） can impact gene function or cause disease. High-throughput RNA sequencing（RNA-seq） has become a powerful technology for transcriptome-wide analysis of AS, but RT-PCR still remains the gold-standard approach for quantifying and validating exon splicing levels. We have developed PrimerSeq, a user-friendly software for systematic design and visualization of RT-PCR primers using RNA-seq data. PrimerSeq incorporates user-provided transcriptome profiles（i.e., RNA-seq data） in the design process, and is particularly useful for largescale quantitative analysis of AS events discovered from RNA-seq experiments. PrimerSeq features a graphical user interface（GUI） that displays the RNA-seq data juxtaposed with the expected RT-PCR results. To enable primer design and visualization on user-provided RNA-seq data and transcript annotations, we have developed PrimerSeq as a stand-alone software that runs on local computers. PrimerSeq is freely available for Windows and Mac OS X along with source code at http：//primerseq.sourceforge.net/. With the growing popularity of RNA-seq for transcriptome studies, we expect PrimerSeq to help bridge the gap between high-throughput RNA-seq discovery of AS events and molecular analysis of candidate events by RT-PCR.

AF1q inhibited T cell attachment to breast cancer cell by attenuating Intracellular Adhesion Molecule-1 expression

Aim: To investigate whether AF1q, overexpressed in metastatic cells compared with the primary tumor cells, plays a pivotal role in breast cancer metastasis. Methods: To investigate whether AF1q has a responsibility in the acquisition of a metastatic phenotype, we performed RNA-sequencing (RNA-Seq) to identify the gene signature and applied the Metacore direct interactions network building algorithm with the top 40 amplicons of RNA-Seq. Results: Most genes were directly linked with intercellular adhesion molecule-1 (ICAM-1). Likewise, we identified that ICAM-1 expression is attenuated in metastatic cells compared to primary tumor cells. Moreover, overexpression of AF1q attenuated ICAM-1 expression, whereas suppression of AF1q elicited the opposite effect. AF1q had an effect on ICAM-1 promoter region and regulated its transcription. Decreased ICAM-1 expression ;affected the attachment of T cells to a breast cancer cell monolayer. We confirmed the finding by performing the analysis on Burkitt's lymphoma. Conclusion: Attenuation of ICAM-1 by AF1q on tumor cells disadvantages host anti-tumor defenses through the trafficking of lymphocytes, which affects tumor progression and metastasis.