Biology subjects

Alexandru-Crivac, C. N.

Publications and source records attributed to Alexandru-Crivac, C. N..

2 recordsLinked to original sources

bioRxiv2024

Large-Scale Comparative Transcriptomic Analysis of CHO Cell Functional Adaptation to Recombinant Monoclonal Antibody Production

To comparatively evaluate cellular constraints on recombinant monoclonal antibody (mAb) production by Chinese Hamster Ovary (CHO) cells, we analysed the transcriptomes of 24 clonally derived CHO cell lines engineered with PiggyBac transposon technology to stably produce four recombinant monoclonal antibodies (mAbs) at varying specific production rates. Fed-batch cultures were sampled at exponential (day 5) and stationary (day 10) phases of culture for analysis by RNA-Seq. Recombinant mRNAs accounted for a large proportion of total mRNA across all clones, and efficient use of heavy chain (HC) mRNA to synthesise recombinant mAb (qP per HC mRNA) varied significantly with respect to both mAb product and cell line. Comparative bioinformatic analyses of CHO transcriptomes focussed on mAb specific production rate and utilised both data-driven and hypothesis-led approaches, specifically (i) production or non-production of recombinant mAb, (ii) changes in the abundance of functional groups of mRNAs abundance with mAb specific production rate and (iii) comparative analysis of informatically-mined gene subsets associated with cellular functions hypothesised to impact recombinant mAb synthesis and secretion. These analyses revealed widespread constitutive and adaptive changes in mRNA abundance associated with mAb production across a variety of cellular functions. Typically, most mechanistically consistent changes in mRNA abundance co-varying with mAb production were evident at the stationary phase sample point. These data revealed both recombinant mAb-specific limitations on cellular synthetic capacity and a generic adaptive strategy used by CHO cells to support high-level mAb production. The latter was achieved by directed and permissive regulation of endoplasmic reticulum and other processes to accommodate increased synthetic flux.

bioengineering↗

bioRxiv2024

Widespread 3' UTR splicing regulates expression of oncogene transcripts in sequence-dependent and independent manners

BackgroundSplicing in 3 untranslated regions (3 UTRs) is generally considered a signal to elicit transcript degradation via nonsense-mediated decay (NMD) due to the presence of an exon junction complex (EJC) downstream of the stop codon. However, 3 UTR intron (3UI)-containing transcripts are widespread and highly expressed in both normal tissues and cancers. ResultsHere we present and characterise a novel transcriptome assembly built from 7897 solid tumour and normal samples from The Cancer Genome Atlas. We identify thousands of 3UI-containing transcript isoforms, many of which are expressed across multiple cancer types. We find that the expression of core NMD component UPF1 negatively correlates with global 3UI splicing between normal samples, however this correlation is lost in colon cancer. We find that 3UIs found exclusively within 3 UTRs (bona-fide 3UIs) are not predominantly NMD-sensitising, unlike introns present in 3 UTRs due to premature termination. We identify HRAS as an example where 3UI splicing rescues the transcript from NMD. Bona-fide, but not premature termination codon (PTC) carrying 3UI-transcripts are spliced more in cancer samples compared to matched normals in the majority of cancer types analysed. In colon cancer, differentially spliced 3UI-containing transcripts are enriched in the canonical Wnt signalling pathway, with CTNNB1 being the most over-spliced in colon cancer compared to normal. We show that manipulating Wnt signalling can further regulate splicing of Wnt component transcript 3 UTRs. ConclusionsOur results indicate that 3 UTR splicing is not a rare occurrence, especially in colon cancer, where 3 UTR splicing regulates transcript expression in EJC-dependent and independent manners.

genomics↗