ArticleStem cell reports2020
N-Terminal Amino Acids Determine KLF4 Protein Stability in 2A Peptide-Linked Polycistronic Reprogramming Constructs.
Article in Stem cell reports, 2020. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 9 papers.
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Who cites it
9 citing papers in PubMed, 17 citations in OpenAlex.
- Multi-gene Co-expression systems inSynthetic and systems biotechnology · 2026Review
- Self-amplifying RNA-based CAR T cell therapy with enhanced duration and multi-genic logic functions.bioRxiv : the preprint server for biology · 2026Article
- The multipurpose cell factory Aspergillus niger can be engineered to produce hydroxylated collagen.Biotechnology for biofuels and bioproducts · 2025Article
- Detailed phenotyping of Tbr1-2A-CreER knock-in mice demonstrates significant impacts on TBR1 protein levels and axon development.Autism research : official journal of the International Society for Autism Research · 2025Article
- LwaCas13a indiscriminately targets the human transcriptome.Wellcome open research · 2025Article
- Synthetic translational coupling element for multiplexed signal processing and cellular control.Nucleic acids research · 2024Article
- An Enhanced Retroviral Vector for Efficient Genetic Manipulation and Selection in Mammalian Cells.Biomolecules · 2024Article
- B1 SINE-binding ZFP266 impedes mouse iPSC generation through suppression of chromatin opening mediated by reprogramming factors.Nature communications · 2023Article
- Structurally-discovered KLF4 variants accelerate and stabilize reprogramming to pluripotency.iScience · 2022Article
Corrections and comments
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Authors and funding
3 authors at 1 institution in 1 country.
Funding
No grant is acknowledged in the PubMed record.
Abstract
A common strategy for multi-protein expression is to link genes by self-cleaving 2A peptide sequences. Yet, little is known how the 2A peptide-derived N-terminal proline or adjacent non-native residues introduced during cDNA cloning affects protein stoichiometry. Polycistronic reprogramming constructs with altered KLF4 protein stoichiometry can influence induced pluripotent stem cell (iPSC) generation. We studied the impact of N-terminal 2A peptide-adjacent residues on the protein stability of two KLF4 isoforms, and assayed their capacity to generate iPSCs. Here, we show that the N-terminal proline remnant of the 2A peptide, alone or in combination with leucine, introduced during polycistronic cloning, destabilizes KLF4 resulting in increased protein degradation, which hinders reprogramming. Interestingly, the addition of charged and hydrophilic amino acids, such as glutamate or lysine stabilizes KLF4, enhancing reprogramming phenotypes. These findings raise awareness that N-terminal modification with 2A peptide-derived proline or additional cloning conventions may affect protein stability within polycistronic constructs.
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Registered trials
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