Difference between revisions of "Part:BBa K3504008"

Revision as of 19:45, 26 October 2020

MiR-126 Binding Site

NOTICE: Parts in our range for this season have been created as a part of our Phase I design of our project. These parts HAVE NOT been tested or characterized in the lab due to COVID-19-related precautionary measures. We have enriched our new parts pages with data from literature and results from our modeling and simulations. If you are intending on using this part or others in our range, please keep in mind these limitations and update these parts with data from your experimentation. Feel free to reach us at: igem.afcm@gmail.com for further inquiries.

Part Description

Binding site for Mir-126. Mir-126 is a short non-coding RNA molecule that function to regulate the expression levels of other genes by several pre-and post-transcription mechanisms.It is a human microRNA that is expressed only in endothelial cells, throughout capillaries as well as larger blood vessels and acts upon various transcripts to control angiogenesis.

Usage

The miRNA-126 has an already established role in angiogenesis and cancer, but recently in the current issue of Nature immunology, it was reported by agudo and colleagues that this miRNA-126 plays a part in survival and function of plasmocytoid dendritic cells (pDCs). These pDCs express Toll-like receptors (TLRs) 7 and 9 inside the cell which once activated results in the production of large quantities of (anti-viral) type I interferon. So this indicated that pDCs have a central role in the immune response. These pDCs are modulated by the miRNA-126 through a pathway (VEGFR2) meaning there is a potential capacity to control pDCs homeostasis and execute new deliberations on the effects on the innate immunity when the vascular endothelial growth factor (VEGF) as a target for therapeutic commitments.In pDCs, miR126 targets a negative regulator of mTOR for degradation. In pDCs, miR126 targets a negative regulator of mTOR for degradation. An overactive mTOR pathway is important for generation of pDCs and for TLR-mediated innate responses to nucleic acids and LV vectors in pDCs. Furthermore, one can take advantage of this observation in vector design to avoid transgene expression in pDCs by incorporation of target sequences into the transcript.Since the miRNA-126 is is highly up regulated in the pDCs we inserted a target site in the circuit vector and by that the miRNA-126 binds to the vector before being phagocytosed by the pDCs. This will cause cleavage of the circuit by the near end by the degradation sequence inserted in the 3' prime of the miRNA126 target site, therefore the circuit would be cut and left open, accordingly this will impair the replication of this vector. This will eventually lead to the unrecognition of the vector by pDCs and by that the other vectors will not be phagocytosed or presented on the surface which prevents the activation of the innate immunity and that makes the circuit safe from the attack of the innate immunity.(1), (2)

Characterization

We characterized this part by structural modelling and simulation of interaction between mir126 and its binding site using mit-126 & ff4 as an input for a NOR gate that could protect our replicon vaccine vectors from being attacked by innate immunity inside dendritic cells and act as a safety switch

Figure 2.This figure shows the sbol format of this composite part as well as way in which a nor gate works.

Figure 1.This figure illustrate the interaction between mir126 and its binding site that showed energy equal to -6.12kcal\mol.

References

1-Agudo, J., Ruzo, A., Tung, N., Salmon, H., Leboeuf, M., Hashimoto, D., . . . Brown, B. (2014, January). The miR-126-VEGFR2 axis controls the innate response to pathogen-associated nucleic acids. Retrieved October 26, 2020, from https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3896265/ 2-Ferretti, C., & La Cava, A. (2014, May). MiR-126, a new modulator of innate immunity. Retrieved October 26, 2020, from https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4085494/ Sequence and Features