EMBO/soda-nel huggingface dataset:
The tyrosine phosphorylation of the lectin receptor-like kinase LORE regulates plant immunity (2019)
Figure 1

Figure 1. The kinase activity of LORE is essential for 3-OH-C10:0 sensing

A. 3-OH-C10:0-induced ROS burst is suppressed by A23 and K252a. Two-week-old Col-0 seedlings were treated with 2μM K252a or 40 μM A23. One hour later, 1 μM 3-OH-C10:0 was used to trigger the ROS burst, and the photo count was recorded by a luminometer. Mock is solvent control. RLU, relative light unit. Values are means ± SD (n=3 biological replicates).

B. A23 and K252a suppress 3-OH-C10:0-induced PTI marker gene expression. Treatments with 3-OH-C10:0 and inhibitors were same as in (A), and the plant leaves were collected for RT-qPCR 4 hrs later. Values are means ± SD (n=3 biological replicates). (ANOVA, P <0.01).

C. MAPK cascade activation was inhibited by A23. Treatments with 3-OH-C10:0 and A23 were same as in (A). The plant leaves were collected at 0, 5, and 10 min for immunoblotting. Activated MAPKs were detected by immunoblotting with anti-phospho-p44/42 MAPK antibody. The corresponding bands are indicated for MPK3 and MPK6. Ponceau staining of Rubisco was used for estimating equal loading in each lane. The experiment was repeated three times with similar results.

D. Tyrosine phosphorylation of LORE in vivo. Two-week-old 35S:LORE-FLAG complementation plants were subjected to immunoprecipitation with anti-FLAG agarose beads. A23 (40 μM) was used to inhibit the protein phosphorylation, and the treatments were same as in (A). Protein abundance of precipitated LORE-FLAG was detected by immunoblotting. The tyrosine phosphorylation was probed by anti-pTyrosine (α-pY) antibody. Similar results were observed in three biological replicates.

E. LORE is an active tyrosine kinase. The α-pY antibody was used to probe tyrosine phosphorylation in recombinant protein MBP-tagged LORE-CD (cytosolic domain). The LORE activation loop conserved K516 was mutated to E, and D613 was mutated to V. The bottom gel showing protein abundance stained by Coomassie Brilliant Blue (CBB).
The tyrosine phosphorylation of the lectin receptor-like kinase LORE regulates plant immunity (2019)
Figure 2

Figure 2. Y600 is a key phosphorylation site of LORE

A. Phosphorylated Y600 of LORE was identified by LC-MS/MS. MBP-tagged LORE-CD was used for in vitro kinase assays. Trypsin digested recombinant proteins were subjected to LC-MS/MS analysis. The doubly-charged peptide with m/z 476.24 matches GLLYLHR of LORE with Y600 phosphorylation. Fragment ions retaining charges in the N and C terminus were denoted by "b" and "y", respectively. pY600 indicated phosphorylated Y600.

B. Y600 mutation reduced LORE kinase activity. MBP-tagged LORE-CD and LORE-Y600F-CD (Y600 tyrosine was substituted with phenylalanine, F) were subjected to kinase reactions. The kinase dead mutant LOREK516E served as a negative control. Autophosphorylated LORE was visualized by autoradiography following SDS-PAGE gel electrophoresis. Protein abundance was stained by CBB.

C. LORE Y600 is phosphorylated in plants. Two-week-old pLORE:LORE-FLAG complementation plant leaves were subjected to immunoprecipitation (IP) with anti-FLAG agarose beads. Y600 specific phosphorylation antibody was used to detect Y600 phosphorylation in LORE. The experiment was repeated three times with similar results.

D. 3-OH-C10:0 induced Y600 phosphorylation. Two-week-old pLORE:LORE-FLAG complementation plants were treated with 1 μM 3-OH-C10:0 at indicated times. Plant leaves were subjected to anti-FLAG IP. Phosphorylation level of Y600 was detected by anti-pY600 antibody. The experiment was repeated three times with similar results.

E. Y600 mutation abolished the ROS burst triggered by 3-OH-C10:0. The transgenic plants were treated with 3-OH-C10:0 and ROS burst was measured. WT(6-1) and WT(10-3) are two independent complementation lines with LORE; Y600F(2-6) and Y600F(7-1) are two independent lines complemented with LOREY600F; and kinase-dead LOREK516E was transformed to lore mutant as a negative control. The 2-3 and 5-2 are two independent lines. Values are means ± SD (n=6 biological replicates). (ANOVA, P <0.01).

F. Y600 is required for growth inhibition by 3-OH-C10:0. The Arabidopsis seedlings were grown on ½ MS containing 2.5 μM 3-OH-C10:0 for six days, and the root length was measured. The experiment was repeated three times with similar results. WT(6-1) and Y600 (2-6) lines were used for the assay. Scale bar=0.4 cm. Values are means ± SD (n=3 biological replicates). (ANOVA, P <0.01).

G. Y600 mutation decreased the disease resistance to Pst DC3000. Plants were spray-inoculated with Pst DC3000 at a concentration of 1×109 cfu/ml, and were subjected to bacterial growth curve analysis at 3 dpi. Values are means ± SD (n=6 biological replicates). (ANOVA, P <0.01).
The tyrosine phosphorylation of the lectin receptor-like kinase LORE regulates plant immunity (2019)
Figure 3

Figure 3. PBLs interacts with LORE in vitro and in vivo

A. LORE interacts with specific RLCKs (PBL34/35/36) in yeast. BD-LORE-CD and AD-PBLs were co-transformed to yeast cells and were screened on synthetic dextrose media lacking leucine and tryptophan (SD/-Leu-Trp). The single yeast colonies were serially diluted onto SD/-Leu-Trp and SD/-Leu-Trp-His (SD media lacking leucine, tryptophan and histidine) to observe yeast cell growth. 3-AT(20 mM) was used to inhibit the autoactivation of LORE. Yeast co-transformed with pGADT7-T+pGBKT7-53(+) served as a positive control, and the yeast co-transformed with pGADT7-T+pGBKT7-lam(-) and AD-PBL39+BD-LORE served as the negative controls. EV, empty vector.

B. LORE interacts with PBLs by MBP pull-down assays. The recombinant GST-LORE-CD and the MBP-PBLs were subjected to MBP pull-down analysis. MBP served as a negative control. The interacting proteins were revealed by anti-GST immunoblotting. The experiment was repeated three times with similar results.

C. Co-immunoprecipitation (Co-IP) of LORE with PBLs. 35S promoter-driven LORE-T7 and PBLs-FLAG were transiently co-expressed in N. benthamiana by agroinfiltration. Plant leaves were immunoprecipated with anti-FLAG beads and the proteins were immunoblotted by anti-FLAG antibody. Co-IP proteins were immunoblotted by anti-T7 antibody. The experiment was repeated three times with similar results.

D. LORE interacts with PBLs, revealed by split-luciferase assays. N. benthamiana leaves were co-infiltrated with Agrobacteria carrying 35S:LORE-nLUC and 35S:cLUC-PBLs. Images of chemiluminescence were obtained by applying 0.5 mM luciferin 36 h post infiltration. Similar results were observed in three biological replicates.
The tyrosine phosphorylation of the lectin receptor-like kinase LORE regulates plant immunity (2019)
Figure 4

Figure 4. PBL triple mutants exhibited compromised PTI triggered by 3-OH-C10:0

A. ROS burst in pbl34/35/36 triple mutant, induced by 3-OH-C10:0. Two-week-old seedlings were treated with 1 μM 3-OH-C10:0, and the ROS burst was measured. Values are means ± SD (n=6 biological replicates). (ANOVA, P <0.01).

B. PTI marker gene expression was reduced in pbl34/35/36 triple mutant in response to 3-OH-C10:0. Treatment is same as in A. Samples were collected for RT-qPCR at 4 h post infiltration. Values are means ± SD (n=3 biological replicates). (Student's t-test, **P< 0.01).

C. pbl34/35/36 triple mutant displayed decreased callose depositions in response to 3-OH-C10:0 treatment. Two-week-old seedlings were sprayed with 1 μM 3-OH-C10:0. The leaves were sampled for callose deposition assays 12 hrs later. The stained callose was counted for 1 mm2 of the leaves. Mock is solvent control. Scale bar=100μm.Values are means ± SD (n=6 leaves). (Student's t-test, **P< 0.01).

D. Root length of pbl34/35/36 triple mutant treated with 3-OH-C10:0. The Arabidopsis seedlings were grown on ½ MS containing 2.5 μM 3-OH-C10:0 for six days, and the root length was measured. Scale bar=0.4cm. Values are means ± SD (n=3 biological replicates). (ANOVA, P <0.01).

E. Disease phenotype of pbl34/35 double, pbl34/35/36 triple mutants and PBL34 overexpression plants. 35S:PBL34-FLAG was introduced to the Col-0 to generate the overexpression lines. The homozygous transgenic plants were used for the assays. The plants were sprayed with Pst DC3000 at a concentration of 1×109 cfu/ml. Plants were subjected to bacterial growth curve analysis at 3 dpi. OE-3# and OE-11# are two independent transgenic lines. Values are means ± SD (n=6 biological replicates). (ANOVA, P <0.01).

F. pbl34/35/36 triple mutant phenotype was rescued by PBL34. PBL34-HA was introduced to the triple mutant driven by its native promoter. Four-week-old homozygous seedlings were inoculated with 5×104 cfu/ml Pst DC3000 , and the leaves were subjected to bacterial growth curve analysis at 3 dpi. CM2-5 and CM7-3 are two independent transgenic lines. Values are means ± SD (n=6 biological replicates). (ANOVA, P <0.01).
The tyrosine phosphorylation of the lectin receptor-like kinase LORE regulates plant immunity (2019)
Figure 6

Figure 6. Pst effector HopAO1 targets LORE for tyrosine dephosphorylation

A. LORE interacts with HopAO1 by MBP pull-down assays. The recombinant GST-HopAO1 and the MBP-LORE-CD were subjected to MBP pull-down analysis. MBP and GST proteins served as the negative controls. The interacting proteins were revealed by anti-GST immunoblotting. The experiment was repeated three times with similar results.

B. LORE interacts with HopAO1 by Co-IP assays. 35S promoter-driven LORE-T7 and HopAO1-FLAG were transiently co-expressed in N. benthamiana by agroinfiltration. Plant leaves were immunoprecipated with anti-FLAG beads and immunoblotted by anti-FLAG. Coimmunoprecipitated proteins were immunoblotted by anti-T7. The experiment was repeated three times with similar results.

C. HopAO1 dephosphorylates LORE Y600 in vitro. The recombinant GST-HopAO1/HopAO1CS and GST-LORE-CD were subjected to in vitro phosphatase assays. Phosphorylation level of Y600 was detected by anti-pY600 immunoblotting. The experiment was repeated three times with similar results.

D. HopAO1 dephosphorylates LORE Y600 in N. benthamiana. LORE-FLAG was transiently co-expressed with GUS-T7, HopAO1-T7 or HopAO1CS-T7 in N. benthamiana leaves mediated by Agrobacteria. The protein was immunoprecipitated and probed by anti-pY600 antibody. The experiment was repeated at least three times with similar results.

E. HopAO1 suppresses 3-OH-C10:0-triggered ROS burst. HopAO1 was ectopically expressed in Col-0 driven by estradiol-inducible promoter. 100μM estradiol was sprayed on Est:HopAO1 plants, and the leaf disc was used for ROS burst assays 12 hrs later. Values are means ± SD (n=6 biological replicates).

F. HopAO1 inhibits 3-OH-C10:0-induced disease resistance. The Est:HopAO1 plants were pretreated with 1 μM 3-OH-C10:0 (C10) and100 μM estradiol, and then the plants were inoculated with 5×104 cfu/ml Pst DC3000 24 hrs later. The bacterial titer was analyzed at 3 dpi. Values are means ± SD (n=6; ANOVA; p < 0.01).

G. Working model: LORE is phosphorylated at tyrosine600 (Y600) at a basal level. In perception of 3-OH-C10:0, LORE Y600 phosphorylation is elevated, which then trans-phosphorylates PBL34 and likely its paralogs. Phosphorylated PBL34 dissociates from LORE, leading to immune activation. In this event, PBL34 kinase activity is required. However, the bacterial effector HopAO1 targets LORE Y600 and dephosphorylate the tyrosine-phosphorylated Y600, which subsequently suppresses immune response in Arabidopsis.
TRIM16 controls assembly and degradation of protein aggregates by modulating the p62-NRF2 axis and autophagy (2018)
Figure 1

Figure 1. TRIM16 is required for assembly of protein aggregates.

A-F (A, D) Representative confocal images of HeLa and TRIM16KO cells treated with (A) MG132 (20 µM, 2h), scale bar: 7.5 µm. (D) H2O2 (200 µM, 2h) and the samples were processed for IF analysis with Ub and p62 antibody. (B, E) Representative confocal images of control siRNA and TRIM16 siRNA transfected cells treated with (B) MG132 (20 µM, 2h), (E) H2O2, (200 µM, 2h), where IF analysis was conducted with Ub and p62 antibodies. (C, F) The graph shows the the percentage of cells with Ub-p62 co-localized dots. Data from ≥10 fields (40X), n=3, Mean ±SD, **p < 0.0003, *p < 0.002 (Student's unpaired t-test).

(G) Western blot (WB) analysis of detergent-soluble and -insoluble fractions of HeLa and TRIM16KO cells treated with MG132 (50 µM, 1h) and probed with indicated antibodies.

(H, I) Densitometric analysis (using ImageJ) of WB normalized with α-Tubulin or Lamin-B1.

(J) Representative confocal images of HeLa and TRIM16KO cells treated with MG132 (20 µM, 2h) and processed for IF analysis with Ub and LC3B antibody.

(K, L) WB analysis of LC3B levels in lysates of HeLa and TRIM16KO cells treated with (K) Bafilomycin A1 (300 nM, 3h) alone or (L) with MG132 (10 µM, 4h) as indicated. L.E, Low exposure; H.E, high exposure.

(M) Representative confocal images HeLa and TRIM16KO cells treated with H2O2 (200 µM, 2h) and processed for IF analysis with Proteostat dye, Ub, and p62 antibody. To show the cellular details, the brightness of TRIM16KO cells images was increased more than control cells in panel P.

(N) Pictorial representation of data presented in Figure 1 and Expanded View Figure 1.

Data information: Unless otherwise stated, scale bar: 10 µm.
TRIM16 controls assembly and degradation of protein aggregates by modulating the p62-NRF2 axis and autophagy (2018)
Figure 2

Figure 2. TRIM16 regulates the p62-KEAP1-NRF2 system.

(A) Western blot analysis of HeLa and TRIM16KO cell lysates probed with indicated antibodies.

(B) Western blot analysis of lysates from HeLa and TRIM16KO cells treated with or without MG132 and probed with indicated antibodies.

(C) Left panel, western blotting with HeLa and TRIM16KO cell lysates probed with indicated antibodies. Right panel, densitometric analysis (mean ±SD) of protein band intensity relative to actin, n=3, ***p < 0.0005 (Student's unpaired t-test).

D-G (D, F) HEK293T cells were either transiently transfected with an empty vector or a Flag-tagged-TRIM16 expressing vector and immunoblotted with Flag or indicated antibodies. L.E, Low exposure; H.E, High Exposure. (E, G) Densitometric analysis (mean ±SD) of protein band intensity relative to actin, n=3, *p < 0.05 (Student's unpaired t-test).

(H) WB analysis of HeLa and TRIM16KO lysates of cells treated with cycloheximide (100 µM) for the indicated period and probed with different antibodies as indicated. The blots of control and TRIM16KO cells are exposed for the equal duration and developed together on the same X-ray film.

(I) Quantification of NRF2, p62, and KEAP1 band intensities relative to actin. n=3, **p < 0.005, *p < 0.05 (Student's unpaired t-test).

(J) Immunoprecipitation (IP) analysis of interaction between endogenous TRIM16 and endogenous p62 in HeLa cell lysates in absence and presence of MG132 (10 µM, 4h). *The panel (J) here, Fig EV2H and Appendix Figure S3H are part of same blots hence the input for TRIM16 is same.

(K) The domain organization map of TRIM16 and deletion constructs cloned as Myc-tagged proteins.

(L, M) IP analysis to map the interaction between p62 and TRIM16 domains in HEK293T cell lysates in (L) absence and (M) presence of MG132 (10 µM, 4h).

(N) IP analysis of the interaction between endogenous TRIM16 and endogenous NRF2 in HeLa cell lysates. Mr, Molecular Weight Marker.

(O) Co-IP analysis to map the interaction between NRF2 and TRIM16 domains in HEK293T cell lysates.

(P) IP analysis of the interaction between endogenous TRIM16 and endogenous KEAP1 in HeLa cell lysates.
TRIM16 controls assembly and degradation of protein aggregates by modulating the p62-NRF2 axis and autophagy (2018)
Figure 3

Figure 3. TRIM16 perturbs the interactions of the p62-KEAP1-NRF2 complex.

(A, B) Co-IP analysis of the interaction between NRF2 and KEAP1 in absence and presence of TRIM16.

(C) Co-IP analysis of the interaction between NRF2 and p62 in absence and presence of TRIM16.

(D) Co-IP analysis of the interaction between KEAP1 and p62 in absence and presence of TRIM16.

(E) Graphical representation of data presented in Figure 2 and 3.
TRIM16 controls assembly and degradation of protein aggregates by modulating the p62-NRF2 axis and autophagy (2018)
Figure 4

Figure 4. TRIM16 regulates ubiquitination of NRF2.

(A-C) Analysis of NRF2 ubiquitination in absence and presence of TRIM16 by Co-IP assays using transiently transfected plasmid constructs as indicated. Two different variants of ubiquitin protein are used, one that can only be ubiquitinated at Lysine 48 residue (HA-K48-UB) and other that can be only ubiquitinated at Lysine 63 residue (HA-K63-UB). All other lysine residues are mutated. L.E, Low exposure; H.E, High Exposure.

(D, E) Analysis of NRF2 ubiquitination in absence and presence of TRIM16 by Ni-NTA pull-down assays using transiently transfected plasmid constructs as indicated. His-tagged ubiquitin which is mutated at all lysines except 63 position is used in these assays.

(F) Analysis of NRF2 ubiquitination in absence and presence of TRIM16 deletion variants by Co-IP assays using transiently transfected plasmid constructs as indicated.

(G) Western blot analysis of lysates from HeLa, TRIM16KO cells and TRIM16KO cells complemented with TRIM16 deletion constructs where cells were treated with MG132 (20 µM, 2h) and the blot is probed with indicated antibodies.

(H) Western blot analysis of lysates from HeLa, TRIM16KO cells and TRIM16KO cells complemented with TRIM16 deletion constructs where cells were treated with MG132 (20 µM, 2h) and the blot is probed with indicated antibodies.
TRIM16 controls assembly and degradation of protein aggregates by modulating the p62-NRF2 axis and autophagy (2018)
Figure 5

Figure 5. TRIM16 regulates stress-induced NRF2 response including ubiquitination of protein aggregates.

(A) Western blot analysis of HeLa and TRIM16KO lysates of cells treated with increasing concentrations of H2O2 as specified and probed with antibodies as indicated.

(B) WB analysis of HeLa and TRIM16KO lysates of cells treated with 400 µM of H2O2 for different durations as indicated and probed with antibodies as shown. The red arrows indicate the time point where the cells were retreated with 400 µM of H2O2. L.E, Low exposure; H.E, High Exposure.

(C) Densitometric analysis of NRF2 band intensity (in panel "B") normalized with actin.

(D) WB analysis of HeLa and TRIM16KO lysates of cells treated with 2.5 µM (2 h) of As2O3.

(E-G) RNA isolated from HeLa and TRIM16KO cells, untreated or treated with MG132 (20 µM, 2h) were subjected to qRT-PCR with primers of genes as indicated. The fold induction in MG132 treated samples is calculated relative to untreated samples. Mean ±SD, n=3, *p < 0.05, **p < 0.005 (Student's unpaired t-test).

(H-J) RNA isolated from control and NRF2 knockdown HeLa cells transfected with control vector or Myc-TRIM16 subjected to qRT-PCR with primers of genes as indicated. Mean ±SD, n=3, **p < 0.005, ***p < 0.0005 (Student's unpaired t-test).

(K) Western blot analysis of HeLa and TRIM16KO lysates of cells treated with h increasing concentrations of H2O2 as specified and probed with antibodies as indicated.

(L) Densitometric analysis of Ub smear intensity normalized with actin. Mean ±SD, n=3, *p < 0.05 (Student's unpaired t-test).

(M) Representative confocal images of control, Ubb and Ube2n siRNA transfected cells treated with H2O2 (200 µM, 2h) and IF was performed with Ub and p62 antibodies. Scale bar: 10 µm.

(N) The graph shows the percentage of cells with protein aggregates. Data from ≥ 10 microscopic fields for each condition (40X), n=2, Mean ±SD.

(O) Western blot analysis of Ubb and Ube2n siRNA transfected cell lysates with antibodies as indicated.

(P) Pictorial representation of results obtained in Figure 1 to 5.
TRIM16 controls assembly and degradation of protein aggregates by modulating the p62-NRF2 axis and autophagy (2018)
Figure 6

Figure 6. NRF2 regulates TRIM16, ubiquitin pathway genes expression, and biogenesis of protein aggregates.

(A) Control or NRF2 siRNA transfected HeLa cells lysates were subjected to western blotting with indicated antibodies.

(B, C) RNA isolated from untreated or MG132-treated (20 µM, 2h) control or NRF2 siRNA transfected cells were subjected to qRT-PCR with primers of genes as indicated. The fold induction in MG132 treated samples is calculated relative to untreated samples. Mean ±SD, n=3, *p < 0.05 (Student's unpaired t-test).

(D, E) WB analysis of control siRNA or NRF2 siRNA transfected lysates of HeLa cells, untreated or treated with MG132 (20 µM, 2h) or H2O2 (200 µM, 2h) as indicated and probed with antibodies as shown. L.E, Low exposure; H.E, High Exposure.

(F, G) Left panels: Representative confocal images of control siRNA and NRF2 siRNA transfected cells treated with (F) MG132 (20 µM, 2h), (G) H2O2 (200 µM, 2h), where IF was conducted with Ub and p62 antibodies. Right panels: fluorescence intensity line tracing corresponding to a white line in zoom panel.

(H) Hela cells, TRIM16KO cells, and TRIM16KO cells complemented with Myc-NRF2 or GFP-NRF2 were treated with MG132 (20 µM, 2h) and subjected to WB analysis with Myc, GFP and indicated antibodies.

(I) Representative confocal images of Hela cells, TRIM16KO cells, and TRIM16KO cells complemented with Myc-NRF2 which were treated with MG132 (20 µM, 2h) and subjected to IF with Ub and p62 antibodies.

(J) Graph represent % of cells with Ub-p62 dots, data from ≥ 10 microscopic fields (40X), n=3, mean ±SD, *p < 0.005 (one-way ANOVA test).

(K) Graphical representation of results obtained in Figure 6.

Data information: Unless otherwise stated, scale bar: 10 µm.
TRIM16 controls assembly and degradation of protein aggregates by modulating the p62-NRF2 axis and autophagy (2018)
Figure 7

Figure 7. TRIM16 mediates selective clearance of aggregated autophagy cargoes.

A-D (A, C) WB analysis of detergent-soluble and -insoluble fractions of HeLa and TRIM16KO cells expressing either (A) Flag-p62 or (C) GFP-polyQ74 probed with Flag or GFP or indicated antibodies. L.E, Low exposure; H.E, High Exposure. (B, D) Densitometric analysis of WB's (shown in panel A and C) normalized with α-Tubulin or Lamin-B1.

(E) Confocal images of GFP-TRIM16 expressing HeLa cells, untreated or treated with MG132 (10 µM, 4h) and/or Bafilomycin A1 (300 nM, 3h) and IF analysis is performed with antibodies as indicated. Right panel graphs: fluorescence intensity line tracing corresponding to a white line in zoom panel. Scale bar: 10 µm.

(F) The graph shows the percentage of cells with TRIM16-Ub or TRIM16-Ub-p62 co-localized dots/aggregates. Data from ≥ 10 microscopic fields, n=2, mean ±SD.

(G) Representative confocal images of GFP-TRIM16 expressing HeLa cells, untreated or treated with MG132 (10 µM, 4h) and/or Bafilomycin A1 (300 nM, 3h) and IF analysis is performed with antibodies as indicated. Right panel graphs: fluorescence intensity line tracing corresponding to a white line in zoom panel. Scale bar: 10 µm.

(H) Co-IP analysis of the interaction between two differently tagged TRIM16 variants.

(I) WB analysis of detergent-soluble and -insoluble DSS-cross-linked fractions of HeLa and TRIM16KO cells, untreated or treated with MG132 (10 µM, 4h) and probed with indicated antibodies.

(J) Co-IP analysis of the interaction between Flag-tagged TRIM16 and Myc-tagged TRIM16 full length and its domain deletion proteins.