Proteasomes in Patient Rectal Cancer and Different Intestine Locations: Where Does Proteasome Pool Change?
Abstract
:Simple Summary
Abstract
1. Introduction
2. Results and Discussion
2.1. Proteasome Activities in Rectal Cancer and Different Intestine Locations In Vitro
2.2. Expression of Proteasome Pool Components and MHC Class I Molecules in Rectal Cancer and Different Intestine Locations
2.3. Proteasome Activities in Native Gel
3. Materials and Methods
3.1. Patient Tissue Samples
3.2. Antibodies and Main Reagents
3.3. Preparation of Clarified Tissue Homogenates
3.4. Detection of Proteasome Activities In Vitro
3.5. Western Blotting
3.6. Detection of Proteasome Activities in Native Gel
3.7. Technical Remarks
- (1)
- We investigated the expression of LMP2 and LMP7 immune subunits, but not MECL1 subunit, since generally MECL1 and LMP2 subunits are mutually required for incorporation into 20S structure, with incorporation of MECL1 depends on LMP2 and incorporation of LMP2 is facilitated by MECL1 [8,9]. Therefore, the change of LMP2 content reflects the change of MECL1 content.
- (2)
- For our investigation, the methods requiring proteasome purification, such as quantitative proteomics and similar techniques, were irrelevant, as different steps of proteasome purification lead to partial or total loss of one or another proteasome form and distortion of in-cell state [35,36]. We used the original method of electrophoresis in native gel, specially developed for crude proteasome fractions, to reflect in-cells state of “alive” proteasomes in tissues most correctly.
3.8. Statistics
4. Conclusions
Supplementary Materials
Author Contributions
Funding
Institutional Review Board Statement
Informed Consent Statement
Data Availability Statement
Acknowledgments
Conflicts of Interest
References
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Factors | Designation |
---|---|
Activities in tumor | (1) |
Activities in adjacent tissue | (2) |
Activities in intestine distal side, 4 cm from tumor | (3) |
Activities in intestine distal side, 2 cm from tumor | (4) |
Activities in intestine proximal side, 8 cm from tumor | (5) |
Activities in intestine proximal side, 15 cm from tumor | (6) |
Activities in intestine proximal side, 20 cm from tumor | (7) |
Activity | Statistical Indicators | ||
---|---|---|---|
df | Fst | p | |
ChTL | 6 | 276.9 | <0.0001 |
CL | 6 | 414.2 | <0.0001 |
LMP7 | 6 | 77.1 | <0.0001 |
LMP2 | 6 | 58.9 | <0.0001 |
PL | Homogenous Groups of ChTL Activity | PL | Homogenous Groups of CL Activity | ||||||||||||||
---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
Fisher LSD Test | Bonferroni Test | Fisher LSD Test | Bonferroni Test | ||||||||||||||
1 | 2 | 3 | 4 | 1 | 2 | 3 | 1 | 2 | 3 | 4 | 5 | 1 | 2 | 3 | 4 | ||
(5) | **** | **** | (4) | **** | **** | ||||||||||||
(3) | **** | **** | **** | (3) | **** | **** | **** | **** | |||||||||
(7) | **** | **** | **** | (6) | **** | **** | **** | **** | **** | ||||||||
(4) | **** | **** | (5) | **** | **** | **** | **** | ||||||||||
(6) | **** | **** | (7) | **** | **** | ||||||||||||
(2) | **** | **** | (2) | **** | **** | ||||||||||||
(1) | **** | **** | (1) | **** | **** |
PL | Homogenous Groups of LMP7 Activity | PL | Homogenous Groups of LMP2 Activity | ||||||||||||
---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
Fisher LSD Test | Bonferroni Test | Fisher LSD Test | Bonferroni Test | ||||||||||||
1 | 2 | 3 | 4 | 1 | 2 | 3 | 1 | 2 | 3 | 4 | 1 | 2 | 3 | ||
(4) | **** | **** | (3) | **** | **** | ||||||||||
(6) | **** | **** | (7) | **** | **** | **** | |||||||||
(3) | **** | **** | **** | (4) | **** | **** | **** | ||||||||
(7) | **** | **** | **** | (5) | **** | **** | **** | ||||||||
(5) | **** | **** | (6) | **** | **** | ||||||||||
(2) | **** | **** | (2) | **** | **** | ||||||||||
(1) | **** | **** | (1) | **** | **** |
PL | C and p Indicators for Different Proteasome Locations | ||||||
---|---|---|---|---|---|---|---|
(1) | (2) | (3) | (4) | (5) | (6) | (7) | |
(1) | 1.0000 | 0.6611 | 0.5466 | 0.6188 | 0.3226 | 0.5058 | 0.4109 |
(2) | p = 0.001 | 1.0000 | 0.6368 | 0.5780 | 0.4690 | 0.4859 | 0.4611 |
(3) | p = 0.010 | p = 0.002 | 1.0000 | 0.8615 | 0.5296 | 0.7880 | 0.6174 |
(4) | p = 0.003 | p = 0.006 | p < 0.001 | 1.0000 | 0.4418 | 0.7709 | 0.6595 |
(5) | p = 0.154 | p = 0.032 | p = 0.014 | p = 0.045 | 1.0000 | 0.4611 | 0.4058 |
(6) | p = 0.019 | p = 0.026 | p < 0.001 | p < 0.001 | p = 0.035 | 1.0000 | 0.8048 |
(7) | p = 0.064 | p = 0.035 | p = 0.003 | p = 0.001 | p = 0.068 | p < 0.001 | 1.0000 |
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Share and Cite
Erokhov, P.A.; Kulikov, A.M.; Karpova, Y.D.; Rodoman, G.V.; Sumedi, I.R.; Goncharov, A.L.; Razbirin, D.V.; Gorelova, V.S.; Sharova, N.P.; Astakhova, T.M. Proteasomes in Patient Rectal Cancer and Different Intestine Locations: Where Does Proteasome Pool Change? Cancers 2021, 13, 1108. https://doi.org/10.3390/cancers13051108
Erokhov PA, Kulikov AM, Karpova YD, Rodoman GV, Sumedi IR, Goncharov AL, Razbirin DV, Gorelova VS, Sharova NP, Astakhova TM. Proteasomes in Patient Rectal Cancer and Different Intestine Locations: Where Does Proteasome Pool Change? Cancers. 2021; 13(5):1108. https://doi.org/10.3390/cancers13051108
Chicago/Turabian StyleErokhov, Pavel A., Alexey M. Kulikov, Yaroslava D. Karpova, Grigory V. Rodoman, Ilia R. Sumedi, Artem L. Goncharov, Dmitry V. Razbirin, Vera S. Gorelova, Natalia P. Sharova, and Tatiana M. Astakhova. 2021. "Proteasomes in Patient Rectal Cancer and Different Intestine Locations: Where Does Proteasome Pool Change?" Cancers 13, no. 5: 1108. https://doi.org/10.3390/cancers13051108