Table 2.
Summary of the results.
 |
Wildtype | R143W | C150R | R162Q (experimental control) | K286* | G337S | A448R | M494V | H119R | V139A | M494fs |
|---|---|---|---|---|---|---|---|---|---|---|---|
| DANN score | – | 0.9993 | 0.9976 | – | 0.9956 | 0.5549 | – | 0.9686 | 0.2877 | 0.4585 | – |
| GERP (NR) | – | 6.0799 | 6.0799 | – | 5.1999 | 5.1999 | – | 5.75 | 3.73 | 4 | – |
| Proven score | – | −6.932 | −10.570 | −3.520 | – | 1.872 | −2.441 | −0.691 | 1.437 | 0.562 | – |
| Proven prediction (cutoff:-2.5) | – | Deleterious | Deleterious | Deleterious | – | Neutral | Neutral | Neutral | Neutral | Neutral | – |
| Mutation taster | – | Disease causing | Disease causing | – | Disease causing | Disease causing | – | Disease causing | Polymorphism | Polymorphism | Disease causing |
| FATHMM | – | Tolerated | Damaging | – | – | Tolerated | – | Tolerated | – | – | – |
| FATHMM-MKL | – | Damaging | Damaging | – | Damaging | Neutral | – | Damaging | Neutral | Neutral | – |
| MetaSVM | – | Tolerated | Damaging | – | – | Tolerated | – | Tolerated | Tolerated | Tolerated | – |
| The expression of full-length IKAROS protein in the nuclear extract in western blotting | Expressed | Expressed | Expressed | Expressed | Not expresseda | Expressed | Expressed | Expressed | Not done | Not done | Not done |
| The expression of full-length IKAROS in wildtype/mutant co-transfected cells in western blotting | Expressed | Expressed | Expressed | Expressed | Expressed | Expressed | Expressed | Expressed | Not done | Not done | Not done |
| DNA/Protein complexes formation in wildtype or mutant IKAROS in EMSA | DNA/Protein bands were observed | DNA/Protein bands were absent | DNA/Protein bands were absent | DNA/Protein bands were absent | DNA/Protein bands were observed | DNA/Protein bands were observed | DNA/Protein bands were observed | DNA/Protein bands were observed | Not done | Not done | Not done |
| DNA/Protein complexes formation in wildtype/mutant co-transfected cells in EMSA | DNA/Protein bands were observed | DNA/Protein bands were observed | DNA/Protein bands were observed | DNA/Protein bands were observed | DNA/Protein bands were observed | DNA/Protein bands were observed | DNA/Protein bands were observed | DNA/Protein bands were observed | Not done | Not done | Not done |
| Pericentromeric-Heterochromatin localization(punctate staining pattern) in wildtype or mutant IKAROS | Punctate pattern observed | Diffused staining pattern was observed | Diffused staining pattern was observed | Diffused staining pattern was observed | Diffused staining pattern was observed | Punctate pattern observed | Punctate pattern observed | Punctate pattern observed | Not done | Not done | Not done |
| Pericentromeric-Heterochromatin localization(punctate staining pattern) in wildtype/mutant co-transfection | Puctate pattern observed | Puctate pattern observed | Puctate pattern observed | Puctate pattern observed | Speckled pattern was formed mainly by the wildtype constructb | Puctate pattern observed | Punctate pattern observed | Punctate pattern observed | Not done | Not done | Not done |
| Molecular defects | – | Loss of function | Loss of function | Loss of function | Loss of function | – | – | – | – | – | – |
a
A band of 31 kDa was observed in the nuclear extract of cells, transfected with this variant. The full-length IKAROS could not be detected in this variant due to the stop-gained mutation leading the truncation in Lys286*.
b
The localization defect of the p.Lys286* mutant, is not terminated with co-expression of wildtype IKAROS, consistent with the loss of its dimerization domain. The localization of the co-transfected wildtype IKAROS remained unaffected in these cells.