In silico analysis and in planta production of recombinant ccl21/IL1β protein and characterization of its in vitro anti-tumor and immunogenic activity
CCL21 has an essential role in anti-tumor immune activity. Epitopes of IL1β have adjuvant activity without causing inflammatory responses. CCR7 and its ligands play a vital role in the immune balance; specifically, in transport of T lymphocytes and antigen-presenting cells such as dendritic cells to...
| Main Authors: | , , , , , |
|---|---|
| Format: | Article |
| Language: | English |
| Published: |
Public Library of Science (PLoS)
2022-01-01
|
| Series: | PLoS ONE |
| Online Access: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9423642/?tool=EBI |
| _version_ | 1798032515226664960 |
|---|---|
| author | Hasan Marashi Maria Beihaghi Masoud Chaboksavar Samad Khaksar Homan Tehrani Ardavan Abiri |
| author_facet | Hasan Marashi Maria Beihaghi Masoud Chaboksavar Samad Khaksar Homan Tehrani Ardavan Abiri |
| author_sort | Hasan Marashi |
| collection | DOAJ |
| description | CCL21 has an essential role in anti-tumor immune activity. Epitopes of IL1β have adjuvant activity without causing inflammatory responses. CCR7 and its ligands play a vital role in the immune balance; specifically, in transport of T lymphocytes and antigen-presenting cells such as dendritic cells to the lymph nodes. This study aimed to produce epitopes of CCL21 and IL1β as a recombinant protein and characterize its in vitro anti-tumor and immunogenic activity. A codon-optimized ccl21/IL1β gene was designed and synthesized from human genes. Stability and binding affinity of CCL21/IL1β protein and CCR7 receptor were examined through in silico analyses. The construct was introduced into N. tabacum to produce this recombinant protein and the structure and function of CCL21/IL1β were examined. Purified protein from transgenic leaves generated a strong signal in SDS PAGE and western blotting assays. FTIR measurement and MALDI-TOF/TOF mass spectrography showed that ccl21/IL-1β was correctly expressed in tobacco plants. Potential activity of purified CCL21/IL1β in stimulating the proliferation and migration of MCF7 cancer cell line was investigated using the wound healing method. The results demonstrated a decrease in survival rate and metastasization of cancer cells in the presence of CCL21/IL1β, and IC50 of CCL21 on MCF7 cells was less than that of non-recombinant protein. Agarose assay on PBMCsCCR7+ showed that CCL21/IL1β has biological activity and there is a distinguishable difference between chemokinetic (CCL21) and chemotactic (FBS) movements. Overall, the results suggest that CCL21/IL1β could be considered an effective adjuvant in future in vivo and clinical tests. |
| first_indexed | 2024-04-11T20:15:14Z |
| format | Article |
| id | doaj.art-de2b02dc77f04843916140342d945f56 |
| institution | Directory Open Access Journal |
| issn | 1932-6203 |
| language | English |
| last_indexed | 2024-04-11T20:15:14Z |
| publishDate | 2022-01-01 |
| publisher | Public Library of Science (PLoS) |
| record_format | Article |
| series | PLoS ONE |
| spelling | doaj.art-de2b02dc77f04843916140342d945f562022-12-22T04:05:00ZengPublic Library of Science (PLoS)PLoS ONE1932-62032022-01-01178In silico analysis and in planta production of recombinant ccl21/IL1β protein and characterization of its in vitro anti-tumor and immunogenic activityHasan MarashiMaria BeihaghiMasoud ChaboksavarSamad KhaksarHoman TehraniArdavan AbiriCCL21 has an essential role in anti-tumor immune activity. Epitopes of IL1β have adjuvant activity without causing inflammatory responses. CCR7 and its ligands play a vital role in the immune balance; specifically, in transport of T lymphocytes and antigen-presenting cells such as dendritic cells to the lymph nodes. This study aimed to produce epitopes of CCL21 and IL1β as a recombinant protein and characterize its in vitro anti-tumor and immunogenic activity. A codon-optimized ccl21/IL1β gene was designed and synthesized from human genes. Stability and binding affinity of CCL21/IL1β protein and CCR7 receptor were examined through in silico analyses. The construct was introduced into N. tabacum to produce this recombinant protein and the structure and function of CCL21/IL1β were examined. Purified protein from transgenic leaves generated a strong signal in SDS PAGE and western blotting assays. FTIR measurement and MALDI-TOF/TOF mass spectrography showed that ccl21/IL-1β was correctly expressed in tobacco plants. Potential activity of purified CCL21/IL1β in stimulating the proliferation and migration of MCF7 cancer cell line was investigated using the wound healing method. The results demonstrated a decrease in survival rate and metastasization of cancer cells in the presence of CCL21/IL1β, and IC50 of CCL21 on MCF7 cells was less than that of non-recombinant protein. Agarose assay on PBMCsCCR7+ showed that CCL21/IL1β has biological activity and there is a distinguishable difference between chemokinetic (CCL21) and chemotactic (FBS) movements. Overall, the results suggest that CCL21/IL1β could be considered an effective adjuvant in future in vivo and clinical tests.https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9423642/?tool=EBI |
| spellingShingle | Hasan Marashi Maria Beihaghi Masoud Chaboksavar Samad Khaksar Homan Tehrani Ardavan Abiri In silico analysis and in planta production of recombinant ccl21/IL1β protein and characterization of its in vitro anti-tumor and immunogenic activity PLoS ONE |
| title | In silico analysis and in planta production of recombinant ccl21/IL1β protein and characterization of its in vitro anti-tumor and immunogenic activity |
| title_full | In silico analysis and in planta production of recombinant ccl21/IL1β protein and characterization of its in vitro anti-tumor and immunogenic activity |
| title_fullStr | In silico analysis and in planta production of recombinant ccl21/IL1β protein and characterization of its in vitro anti-tumor and immunogenic activity |
| title_full_unstemmed | In silico analysis and in planta production of recombinant ccl21/IL1β protein and characterization of its in vitro anti-tumor and immunogenic activity |
| title_short | In silico analysis and in planta production of recombinant ccl21/IL1β protein and characterization of its in vitro anti-tumor and immunogenic activity |
| title_sort | in silico analysis and in planta production of recombinant ccl21 il1β protein and characterization of its in vitro anti tumor and immunogenic activity |
| url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9423642/?tool=EBI |
| work_keys_str_mv | AT hasanmarashi insilicoanalysisandinplantaproductionofrecombinantccl21il1bproteinandcharacterizationofitsinvitroantitumorandimmunogenicactivity AT mariabeihaghi insilicoanalysisandinplantaproductionofrecombinantccl21il1bproteinandcharacterizationofitsinvitroantitumorandimmunogenicactivity AT masoudchaboksavar insilicoanalysisandinplantaproductionofrecombinantccl21il1bproteinandcharacterizationofitsinvitroantitumorandimmunogenicactivity AT samadkhaksar insilicoanalysisandinplantaproductionofrecombinantccl21il1bproteinandcharacterizationofitsinvitroantitumorandimmunogenicactivity AT homantehrani insilicoanalysisandinplantaproductionofrecombinantccl21il1bproteinandcharacterizationofitsinvitroantitumorandimmunogenicactivity AT ardavanabiri insilicoanalysisandinplantaproductionofrecombinantccl21il1bproteinandcharacterizationofitsinvitroantitumorandimmunogenicactivity |