Unknown,Transcriptomics,Genomics,Proteomics

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Molecular characterization of hybridoma subclones spontaneously switching at high frequencies in vitro

ABSTRACT: This is a comparative microarray analysis of 36-65 hybridomas switching at normal low frequencies in vitro vs. selected variants spontaneously switching at 100 fold higher frequencies in vitro. This analysis reveals differential gene expression of 2091 unique probe sets, 1637 of them corresponded to known genes (647 of which were up-regulated and 990 down-regulated in the variants spontaneously switching at higher frequencies), including genes known to be involved in regular class switching (such as Cd40, Lig4, H2afx,Ung, Xrcc4, Tcfe2a, Id2, Apex1, Bcl6 and Prdm1) as well as genes involved in processes known to be necessary for regular class switch recombination (CSR), such as transcription, response to DNA damage and DNA repair, DNA binding, chromatin modifications, etc. Two subclones of each hybridoma switching at low (L) and high (H) frequencies were analyzed.

ORGANISM(S): Mus musculus

SUBMITTER: Jiri Zavadil

PROVIDER: E-GEOD-15357 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Project description:Changes in gravitational force such as that experienced by astronauts during space flight induce a redistribution of fluids from the caudad to the cephalad portion of the body together with an elimination of normal head-to-foot hydrostatic pressure gradients. To assess brain gene profile changes associated with microgravity and fluid shift, a large-scale analysis of mRNA expression levels was performed in the brains of 2 weeks control and hindlimb-unloaded (HU) mice using cDNA microarrays. Although to different extent, all functional categories displayed significantly regulated genes indicating that considerable transcriptomic alterations are induced by HU. Interestingly, the TIC class (transport of small molecules and ions into the cells) had the highest percentage of up-regulated genes, while the most down-regulated genes were those of the JAE class (Cell junction, Adhesion, Extracellular Matrix). TIC genes comprised 16% of those whose expression was altered, including sodium channel, nonvoltage-gated 1 beta (Scnn1b), glutamate receptor (Grin1), voltage-dependent anion channel 1 (Vdac1), calcium channel beta 3 subunit (Cacnb3) and others. The analysis performed by GeneMAPP revealed several altered protein classes and functional pathways such as blood coagulation and immune response, learning and memory, ion channels and cell junction. In particular, data indicate that HU causes an alteration in hemostasis which resolves in a shift toward a more hyper-coagulative state with an increased risk of venous thrombosis. Furthermore, HU treatment seems to impact on key steps of synaptic plasticity and learning processes. We used brains of four control (C1, C2, C3, C4) and four tail-suspended hindlimb-unloaded (E1, E2, E3, E4) mice to ensure statistical relevance of the study. 60 ug of total RNA extracted in TRIzol from each brain was reversed transcribed into labeled cDNAs using fluorescent Cy5 or Cy3-dUTP (Amersham Biosciences, NJ). Differently labeled cDNAs obtained from a pair of biological replicas were co-hybridized overnight at 50C with a microscope slide spotted with ~ 27,000 mouse cDNA sequences produced by the Microarray Core Facility of the Albert Einstein College of Medicine in the “multiple yellow” design (i.e.: C1C2, C3C4, E1E2, E3E4), described in Iacobas DA, Fan C, Iacobas S et al., Transcriptomic changes in developing kidney exposed to chronic hypoxia. Biochem Biophys Res Commun. 2006 349(1):329-38).

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Molecular characterization of hybridoma subclones spontaneously switching at high frequencies in vitro

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