Phenotypic Mutation 'Daffy' (pdf version)
AlleleDaffy
Mutation Type missense
Chromosome16
Coordinate15,647,561 bp (GRCm39)
Base Change A ⇒ T (forward strand)
Gene Prkdc
Gene Name protein kinase, DNA activated, catalytic polypeptide
Synonym(s) slip, DNA-PK, XRCC7, DNAPDcs, DOXNPH, dxnph, DNA-PKcs
Chromosomal Location 15,455,730-15,660,099 bp (+) (GRCm39)
MGI Phenotype FUNCTION: [Summary is not available for the mouse gene. This summary is for the human ortholog.] This gene encodes the catalytic subunit of the DNA-dependent protein kinase (DNA-PK). It functions with the Ku70/Ku80 heterodimer protein in DNA double strand break repair and recombination. The protein encoded is a member of the PI3/PI4-kinase family.[provided by RefSeq, Jul 2010]
PHENOTYPE: Mutations at this locus effect genome stability, radiation sensitivity and DNA repair. Nonsense (scid) and null homozygotes have severe combined immunodeficiency. A BALB/c variant allele reduces enzyme activity and predisposes to breast cancer. [provided by MGI curators]
Accession Number

NCBI RefSeq: NM_011159; MGI:104779

MappedYes 
Amino Acid Change Isoleucine changed to Phenylalanine
Institutional SourceBeutler Lab
Gene Model predicted gene model for protein(s): [ENSMUSP00000023352]
AlphaFold no structure available at present
SMART Domains Protein: ENSMUSP00000023352
Gene: ENSMUSG00000022672
AA Change: I3662F

DomainStartEndE-ValueType
low complexity region 125 138 N/A INTRINSIC
low complexity region 1253 1263 N/A INTRINSIC
low complexity region 1508 1522 N/A INTRINSIC
NUC194 1810 2206 2.37e-246 SMART
SCOP:d1gw5a_ 2210 2493 5e-3 SMART
low complexity region 2669 2681 N/A INTRINSIC
low complexity region 2841 2855 N/A INTRINSIC
Pfam:FAT 3024 3470 8.2e-75 PFAM
PI3Kc 3749 4068 3.67e-86 SMART
FATC 4096 4128 1.57e-9 SMART
Predicted Effect possibly damaging

PolyPhen 2 Score 0.861 (Sensitivity: 0.83; Specificity: 0.93)
(Using ENSMUST00000023352)
Meta Mutation Damage Score 0.1079 question?
Is this an essential gene? Probably essential (E-score: 0.953) question?
Phenotypic Category Unknown
Candidate Explorer Status loading ...
Single pedigree
Linkage Analysis Data
Penetrance  
Alleles Listed at MGI

All Mutations and Alleles(43) : Chemically induced (ENU)(15) Endonuclease-mediated(3) Gene trapped(17) Spontaneous(2) Targeted(5) Transgenic(1)

Lab Alleles
AlleleSourceChrCoordTypePredicted EffectPPH Score
IGL00157:Prkdc APN 16 15515090 missense probably damaging 1.00
IGL00225:Prkdc APN 16 15627508 missense possibly damaging 0.64
IGL00481:Prkdc APN 16 15608330 missense probably benign 0.41
IGL00488:Prkdc APN 16 15593711 splice site probably null
IGL00489:Prkdc APN 16 15617790 missense possibly damaging 0.51
IGL00579:Prkdc APN 16 15482103 missense probably damaging 1.00
IGL00587:Prkdc APN 16 15470222 splice site probably benign
IGL00666:Prkdc APN 16 15554699 missense probably damaging 1.00
IGL00675:Prkdc APN 16 15605022 missense probably benign 0.05
IGL00708:Prkdc APN 16 15597290 missense probably damaging 0.97
IGL00725:Prkdc APN 16 15634503 missense probably benign 0.10
IGL00818:Prkdc APN 16 15577618 missense possibly damaging 0.92
IGL00917:Prkdc APN 16 15557428 missense probably damaging 0.98
IGL00990:Prkdc APN 16 15519979 missense probably benign 0.03
IGL01126:Prkdc APN 16 15487185 missense probably benign 0.01
IGL01141:Prkdc APN 16 15544568 missense probably damaging 0.99
IGL01306:Prkdc APN 16 15485595 missense possibly damaging 0.67
IGL01326:Prkdc APN 16 15647556 missense probably benign
IGL01335:Prkdc APN 16 15634760 critical splice donor site probably null
IGL01419:Prkdc APN 16 15653030 missense probably damaging 1.00
IGL01434:Prkdc APN 16 15531451 missense probably benign 0.00
IGL01554:Prkdc APN 16 15470166 missense probably benign 0.05
IGL01671:Prkdc APN 16 15485609 missense possibly damaging 0.90
IGL01871:Prkdc APN 16 15600951 missense probably benign 0.00
IGL01874:Prkdc APN 16 15552858 missense possibly damaging 0.89
IGL01930:Prkdc APN 16 15516751 missense probably damaging 1.00
IGL01984:Prkdc APN 16 15526643 missense probably benign
IGL02121:Prkdc APN 16 15535048 missense probably benign 0.18
IGL02152:Prkdc APN 16 15487149 missense probably benign 0.15
IGL02172:Prkdc APN 16 15627623 missense probably benign 0.10
IGL02336:Prkdc APN 16 15603843 missense probably benign 0.01
IGL02336:Prkdc APN 16 15603842 missense possibly damaging 0.47
IGL02393:Prkdc APN 16 15634622 missense probably benign 0.42
IGL02406:Prkdc APN 16 15488399 missense probably benign 0.00
IGL02500:Prkdc APN 16 15532146 critical splice donor site probably null
IGL02568:Prkdc APN 16 15544406 missense probably damaging 0.98
IGL02579:Prkdc APN 16 15488465 missense possibly damaging 0.83
IGL02652:Prkdc APN 16 15600951 missense probably benign 0.00
IGL02661:Prkdc APN 16 15587689 missense possibly damaging 0.92
IGL02685:Prkdc APN 16 15653907 missense possibly damaging 0.61
IGL02741:Prkdc APN 16 15570590 splice site probably benign
IGL02803:Prkdc APN 16 15651530 splice site probably benign
IGL02866:Prkdc APN 16 15649191 missense probably damaging 1.00
IGL02882:Prkdc APN 16 15469383 nonsense probably null
IGL02989:Prkdc APN 16 15617880 missense possibly damaging 0.67
IGL03053:Prkdc APN 16 15652030 missense probably benign 0.02
IGL03071:Prkdc APN 16 15617848 missense probably benign 0.01
IGL03091:Prkdc APN 16 15523174 splice site probably benign
IGL03100:Prkdc APN 16 15531499 missense probably benign 0.08
IGL03128:Prkdc APN 16 15518608 splice site probably benign
IGL03168:Prkdc APN 16 15652030 missense probably benign 0.02
IGL03204:Prkdc APN 16 15587665 missense probably benign 0.01
IGL03390:Prkdc APN 16 15488490 nonsense probably null
anhimid UTSW 16 15543325 critical splice donor site probably null
anhinga UTSW 16 15526796 critical splice donor site probably null
Bushtit UTSW 16 15570628 missense probably damaging 0.97
clover UTSW 16 15520020 splice site probably benign
crackle UTSW 16 15603914 critical splice donor site probably null
darter UTSW 16 15591477 missense possibly damaging 0.93
Elmer_fudd UTSW 16 15625922 missense probably benign 0.01
envenomation UTSW 16 15653091 nonsense probably null
hobgoblin UTSW 16 15633850 missense probably damaging 1.00
Incubus UTSW 16 15490191 missense probably damaging 1.00
liming UTSW 16 15570693 nonsense probably null
newt UTSW 16 15545590 missense probably benign 0.04
ornithorhynchus UTSW 16 15634523 critical splice donor site probably null
primitive UTSW 16 15653022 frame shift probably null
roadrunner UTSW 16 15651751 missense probably damaging 1.00
Schreier UTSW 16 15488392 missense probably benign 0.00
screamer UTSW 16 15649146 nonsense probably null
Screamer10 UTSW 16 15585889 missense probably damaging 0.98
screamer2 UTSW 16 15470416 critical splice donor site probably null
screamer3 UTSW 16 15558196 critical splice donor site probably null
screamer4 UTSW 16 15600943 missense probably benign 0.00
screamer5 UTSW 16 15505268 missense probably benign
screamer6 UTSW 16 15577469 missense probably damaging 1.00
screamer7 UTSW 16 15472681 splice site probably null
Screamer8 UTSW 16 15537297 missense probably benign 0.00
Screamer9 UTSW 16 15552786 missense probably benign 0.01
Tweetie UTSW 16 15535665 missense probably damaging 1.00
updock UTSW 16 15612958 missense probably benign
ANU23:Prkdc UTSW 16 15485595 missense possibly damaging 0.67
R0008:Prkdc UTSW 16 15526565 splice site probably benign
R0018:Prkdc UTSW 16 15544406 missense probably benign 0.03
R0018:Prkdc UTSW 16 15544406 missense probably benign 0.03
R0069:Prkdc UTSW 16 15544368 missense probably benign 0.03
R0125:Prkdc UTSW 16 15516871 missense probably damaging 0.98
R0131:Prkdc UTSW 16 15531517 missense probably benign 0.09
R0131:Prkdc UTSW 16 15531517 missense probably benign 0.09
R0132:Prkdc UTSW 16 15531517 missense probably benign 0.09
R0137:Prkdc UTSW 16 15558196 critical splice donor site probably null
R0334:Prkdc UTSW 16 15554663 missense probably benign 0.00
R0373:Prkdc UTSW 16 15609791 missense probably damaging 1.00
R0485:Prkdc UTSW 16 15651604 missense probably damaging 0.97
R0511:Prkdc UTSW 16 15649146 nonsense probably null
R0538:Prkdc UTSW 16 15651652 missense probably damaging 1.00
R0595:Prkdc UTSW 16 15625952 missense probably damaging 1.00
R0607:Prkdc UTSW 16 15589921 missense probably damaging 0.98
R0616:Prkdc UTSW 16 15508271 missense probably damaging 1.00
R0630:Prkdc UTSW 16 15628665 missense probably damaging 1.00
R0694:Prkdc UTSW 16 15586501 missense probably damaging 1.00
R0702:Prkdc UTSW 16 15603835 missense possibly damaging 0.95
R0965:Prkdc UTSW 16 15647580 missense probably benign
R1027:Prkdc UTSW 16 15468576 missense possibly damaging 0.80
R1029:Prkdc UTSW 16 15472613 splice site probably benign
R1033:Prkdc UTSW 16 15585815 missense probably damaging 1.00
R1067:Prkdc UTSW 16 15570646 missense probably damaging 0.99
R1116:Prkdc UTSW 16 15600943 missense probably benign 0.00
R1187:Prkdc UTSW 16 15577610 missense probably damaging 0.98
R1226:Prkdc UTSW 16 15491861 missense possibly damaging 0.80
R1279:Prkdc UTSW 16 15508146 missense probably damaging 1.00
R1304:Prkdc UTSW 16 15577587 missense probably damaging 0.99
R1314:Prkdc UTSW 16 15482091 missense possibly damaging 0.68
R1351:Prkdc UTSW 16 15485564 missense possibly damaging 0.62
R1509:Prkdc UTSW 16 15549430 missense probably damaging 1.00
R1512:Prkdc UTSW 16 15505268 missense probably benign
R1531:Prkdc UTSW 16 15589970 missense probably benign 0.01
R1579:Prkdc UTSW 16 15493192 missense probably benign 0.00
R1669:Prkdc UTSW 16 15551922 missense probably damaging 1.00
R1682:Prkdc UTSW 16 15494853 missense probably benign 0.19
R1713:Prkdc UTSW 16 15612958 missense probably benign
R1762:Prkdc UTSW 16 15455825 missense probably benign
R1789:Prkdc UTSW 16 15557388 missense probably damaging 1.00
R1822:Prkdc UTSW 16 15577469 missense probably damaging 1.00
R1848:Prkdc UTSW 16 15625922 missense probably benign 0.01
R1887:Prkdc UTSW 16 15647499 missense probably benign 0.00
R1891:Prkdc UTSW 16 15543300 missense probably benign 0.02
R1921:Prkdc UTSW 16 15532079 missense possibly damaging 0.80
R1922:Prkdc UTSW 16 15532130 missense probably benign 0.00
R1929:Prkdc UTSW 16 15472681 splice site probably null
R1939:Prkdc UTSW 16 15653777 missense possibly damaging 0.95
R2021:Prkdc UTSW 16 15494873 missense probably benign 0.00
R2033:Prkdc UTSW 16 15505216 splice site probably benign
R2056:Prkdc UTSW 16 15545469 missense probably benign 0.03
R2057:Prkdc UTSW 16 15545469 missense probably benign 0.03
R2058:Prkdc UTSW 16 15545469 missense probably benign 0.03
R2082:Prkdc UTSW 16 15533827 missense probably damaging 1.00
R2109:Prkdc UTSW 16 15505254 missense probably benign 0.01
R2124:Prkdc UTSW 16 15537297 missense probably benign 0.00
R2164:Prkdc UTSW 16 15523071 missense probably damaging 1.00
R2174:Prkdc UTSW 16 15552786 missense probably benign 0.01
R2191:Prkdc UTSW 16 15516688 missense probably damaging 1.00
R2270:Prkdc UTSW 16 15472681 splice site probably null
R2271:Prkdc UTSW 16 15472681 splice site probably null
R2272:Prkdc UTSW 16 15472681 splice site probably null
R2356:Prkdc UTSW 16 15502068 missense probably benign
R2852:Prkdc UTSW 16 15470416 critical splice donor site probably null
R3115:Prkdc UTSW 16 15482222 missense probably benign 0.01
R3116:Prkdc UTSW 16 15482222 missense probably benign 0.01
R3499:Prkdc UTSW 16 15585889 missense probably damaging 0.98
R3687:Prkdc UTSW 16 15617831 missense probably benign
R3834:Prkdc UTSW 16 15609810 missense probably damaging 1.00
R3835:Prkdc UTSW 16 15609810 missense probably damaging 1.00
R3961:Prkdc UTSW 16 15647475 splice site probably null
R4151:Prkdc UTSW 16 15634637 missense probably benign
R4233:Prkdc UTSW 16 15653783 missense probably benign 0.11
R4281:Prkdc UTSW 16 15623963 splice site probably null
R4296:Prkdc UTSW 16 15555769 missense probably damaging 0.99
R4344:Prkdc UTSW 16 15585886 missense probably damaging 0.98
R4424:Prkdc UTSW 16 15653946 missense probably damaging 1.00
R4424:Prkdc UTSW 16 15591603 missense probably damaging 0.98
R4497:Prkdc UTSW 16 15518517 missense probably benign 0.43
R4549:Prkdc UTSW 16 15554734 missense possibly damaging 0.89
R4594:Prkdc UTSW 16 15585830 missense possibly damaging 0.64
R4603:Prkdc UTSW 16 15628688 missense probably damaging 0.98
R4615:Prkdc UTSW 16 15480938 missense probably damaging 0.99
R4648:Prkdc UTSW 16 15634638 missense probably benign 0.05
R4662:Prkdc UTSW 16 15551916 missense probably damaging 1.00
R4680:Prkdc UTSW 16 15589894 missense probably benign 0.00
R4700:Prkdc UTSW 16 15519976 missense probably damaging 1.00
R4716:Prkdc UTSW 16 15628701 missense probably benign 0.32
R4720:Prkdc UTSW 16 15485579 missense probably benign
R4785:Prkdc UTSW 16 15466840 missense probably benign 0.21
R4822:Prkdc UTSW 16 15468576 missense possibly damaging 0.80
R4829:Prkdc UTSW 16 15519939 missense possibly damaging 0.80
R4981:Prkdc UTSW 16 15496173 missense probably damaging 1.00
R4989:Prkdc UTSW 16 15491861 missense possibly damaging 0.80
R5059:Prkdc UTSW 16 15655882 missense probably damaging 1.00
R5074:Prkdc UTSW 16 15589912 missense probably damaging 1.00
R5115:Prkdc UTSW 16 15608444 missense probably benign
R5151:Prkdc UTSW 16 15533899 missense probably damaging 1.00
R5165:Prkdc UTSW 16 15496136 missense probably damaging 1.00
R5215:Prkdc UTSW 16 15589985 missense possibly damaging 0.64
R5270:Prkdc UTSW 16 15552819 missense probably damaging 1.00
R5278:Prkdc UTSW 16 15532838 missense probably damaging 1.00
R5351:Prkdc UTSW 16 15649176 missense probably benign 0.03
R5416:Prkdc UTSW 16 15623814 missense probably damaging 1.00
R5418:Prkdc UTSW 16 15612961 missense probably benign 0.20
R5437:Prkdc UTSW 16 15587739 missense possibly damaging 0.46
R5452:Prkdc UTSW 16 15586501 missense possibly damaging 0.96
R5518:Prkdc UTSW 16 15496172 missense probably damaging 1.00
R5538:Prkdc UTSW 16 15469333 missense probably damaging 1.00
R5589:Prkdc UTSW 16 15524655 missense probably benign 0.02
R5618:Prkdc UTSW 16 15627476 missense probably damaging 1.00
R5640:Prkdc UTSW 16 15647633 missense possibly damaging 0.86
R5661:Prkdc UTSW 16 15628634 missense possibly damaging 0.81
R5771:Prkdc UTSW 16 15482097 missense probably damaging 1.00
R5772:Prkdc UTSW 16 15597252 missense possibly damaging 0.49
R5783:Prkdc UTSW 16 15535665 missense probably damaging 1.00
R5792:Prkdc UTSW 16 15634616 missense probably damaging 1.00
R5797:Prkdc UTSW 16 15555698 nonsense probably null
R5826:Prkdc UTSW 16 15551962 missense probably benign
R5883:Prkdc UTSW 16 15533778 missense probably benign
R5895:Prkdc UTSW 16 15570693 nonsense probably null
R5998:Prkdc UTSW 16 15601021 missense probably damaging 1.00
R6000:Prkdc UTSW 16 15647561 missense possibly damaging 0.86
R6120:Prkdc UTSW 16 15557335 missense probably benign 0.00
R6145:Prkdc UTSW 16 15589937 missense probably damaging 1.00
R6209:Prkdc UTSW 16 15608456 missense probably damaging 1.00
R6293:Prkdc UTSW 16 15605019 missense probably benign 0.00
R6321:Prkdc UTSW 16 15532783 missense probably benign
R6376:Prkdc UTSW 16 15587749 missense probably benign 0.06
R6387:Prkdc UTSW 16 15516679 missense probably benign 0.01
R6406:Prkdc UTSW 16 15535665 missense probably damaging 1.00
R6469:Prkdc UTSW 16 15612939 missense probably benign 0.10
R6486:Prkdc UTSW 16 15570628 missense probably damaging 0.97
R6665:Prkdc UTSW 16 15603914 critical splice donor site probably null
R6703:Prkdc UTSW 16 15488392 missense probably benign 0.00
R6774:Prkdc UTSW 16 15543325 critical splice donor site probably null
R6854:Prkdc UTSW 16 15469402 missense probably damaging 1.00
R6878:Prkdc UTSW 16 15594936 missense probably benign 0.31
R6882:Prkdc UTSW 16 15626020 missense probably benign 0.33
R6882:Prkdc UTSW 16 15601127 critical splice donor site probably null
R6949:Prkdc UTSW 16 15617853 missense probably benign
R6950:Prkdc UTSW 16 15633850 missense probably damaging 1.00
R7019:Prkdc UTSW 16 15587830 missense probably benign 0.00
R7064:Prkdc UTSW 16 15608317 missense probably benign 0.00
R7097:Prkdc UTSW 16 15507207 missense probably damaging 1.00
R7201:Prkdc UTSW 16 15516667 missense probably benign 0.12
R7235:Prkdc UTSW 16 15532127 missense probably benign
R7283:Prkdc UTSW 16 15535628 missense probably benign 0.00
R7401:Prkdc UTSW 16 15466602 missense probably damaging 1.00
R7525:Prkdc UTSW 16 15490191 missense probably damaging 1.00
R7647:Prkdc UTSW 16 15555807 missense probably damaging 1.00
R7679:Prkdc UTSW 16 15649183 missense probably damaging 1.00
R7803:Prkdc UTSW 16 15623960 missense probably null 0.05
R7858:Prkdc UTSW 16 15507141 missense probably benign 0.11
R7872:Prkdc UTSW 16 15532870 missense probably benign 0.05
R7896:Prkdc UTSW 16 15526767 missense probably damaging 0.97
R8032:Prkdc UTSW 16 15597315 missense probably benign 0.00
R8055:Prkdc UTSW 16 15634749 missense probably benign 0.09
R8153:Prkdc UTSW 16 15482108 missense probably damaging 1.00
R8281:Prkdc UTSW 16 15523117 missense probably damaging 1.00
R8302:Prkdc UTSW 16 15653946 missense probably damaging 1.00
R8322:Prkdc UTSW 16 15532005 splice site probably benign
R8401:Prkdc UTSW 16 15591477 missense possibly damaging 0.93
R8440:Prkdc UTSW 16 15653022 frame shift probably null
R8458:Prkdc UTSW 16 15608540 critical splice donor site probably null
R8472:Prkdc UTSW 16 15469400 missense probably damaging 1.00
R8478:Prkdc UTSW 16 15466788 missense probably benign 0.00
R8515:Prkdc UTSW 16 15482232 missense probably damaging 1.00
R8546:Prkdc UTSW 16 15480899 missense probably damaging 1.00
R8678:Prkdc UTSW 16 15526796 critical splice donor site probably null
R8739:Prkdc UTSW 16 15626068 missense probably benign 0.01
R8749:Prkdc UTSW 16 15601029 missense possibly damaging 0.85
R8836:Prkdc UTSW 16 15545523 missense probably damaging 1.00
R8904:Prkdc UTSW 16 15545590 missense probably benign 0.04
R8952:Prkdc UTSW 16 15491624 intron probably benign
R8971:Prkdc UTSW 16 15493229 missense probably null 0.99
R8974:Prkdc UTSW 16 15617726 splice site probably null
R9052:Prkdc UTSW 16 15508160 missense probably benign 0.05
R9069:Prkdc UTSW 16 15653091 nonsense probably null
R9200:Prkdc UTSW 16 15523153 missense probably damaging 1.00
R9235:Prkdc UTSW 16 15651751 missense probably damaging 1.00
R9278:Prkdc UTSW 16 15634523 critical splice donor site probably null
R9309:Prkdc UTSW 16 15526792 nonsense probably null
R9386:Prkdc UTSW 16 15496136 missense probably damaging 0.99
R9452:Prkdc UTSW 16 15485465 missense possibly damaging 0.90
R9500:Prkdc UTSW 16 15657079 missense possibly damaging 0.76
R9608:Prkdc UTSW 16 15548335 missense probably damaging 1.00
R9608:Prkdc UTSW 16 15548334 missense possibly damaging 0.96
R9636:Prkdc UTSW 16 15548341 missense probably benign 0.19
R9656:Prkdc UTSW 16 15617818 missense probably benign 0.00
R9674:Prkdc UTSW 16 15533819 missense probably damaging 0.98
R9760:Prkdc UTSW 16 15657044 nonsense probably null
X0023:Prkdc UTSW 16 15558142 missense probably benign
Z1176:Prkdc UTSW 16 15505286 nonsense probably null
Mode of Inheritance Unknown
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Last Updated 2020-07-29 6:45 PM by External Program
Record Created 2019-01-22 2:53 PM by Bruce Beutler
Record Posted 2019-02-01
Phenotypic Description

Figure 1. Daffy mice exhibit increased body weights compared to wild-type littermates. Scaled weights are shown. Abbreviations: WT, wild-type; REF, homozygous reference mice; HET, heterozygous variant mice; VAR, homozygous variant mice. Mean (μ) and standard deviation (σ) are indicated.

Figure 2. Daffy mice exhibit increased CD44 expression on peripheral blood CD4 T cells. Flow cytometric analysis of peripheral blood was utilized to determine CD44 mean fluorescence intensity. Normalized data are shown. Abbreviations: WT, wild-type; REF, homozygous reference mice; HET, heterozygous variant mice; VAR, homozygous variant mice. Mean (μ) and standard deviation (σ) are indicated.

The daffy phenotype was identified among G3 mice of the pedigree R6000, some of which showed increased body weights compared to wild-type littermates (Figure 1). Some mice also showed increased CD44 expression on peripheral blood CD4 T cells (Figure 2).

Nature of Mutation

Figure 3. Linkage mapping of the increased body weights using a recessive model of inheritance. Manhattan plot shows -log10 P values (Y-axis) plotted against the chromosome positions of 67 mutations (X-axis) identified in the G1 male of pedigree R6000. Weight data are shown for single locus linkage analysis without consideration of G2 dam identity. Horizontal pink and red lines represent thresholds of P = 0.05, and the threshold for P = 0.05 after applying Bonferroni correction, respectively.

Whole exome HiSeq sequencing of the G1 grandsire identified 67 mutations. Both of the above anomalies were linked to mutations in two genes on chromosome 16: Prkdc and Lamp3. The mutation in Prkdc was presumed causative as the daffy phenotypes mimic that of other Prkdc mutant alleles (see MGI). The Prkdc mutation is an A to T transversion at base pair 15,829,697 (v38) on chromosome 16, or base pair 192,255 in the GenBank genomic region NC_000082 for Prkdc. The strongest association was found with a recessive model of inheritance to the body weight phenotype, wherein two variant homozygotes departed phenotypically from nine homozygous reference mice and 11 heterozygotes with a P value of 0.000211  (Figure 3). A semidominant effect was observed in the CD44+ CD4 mean fluorescence intensity assay.  

The mutation corresponds to residue 11,007 in the mRNA sequence NM_011159 within exon 77 of 86 total exons.

10992 GATGACTTTTGTAAGATCACTGGCTCGCTACTT

3657  -D--D--F--C--K--I--T--G--S--L--L-

The mutated nucleotide is indicated in red.  The mutation results in an isoleucine to phenylalanine substitution at position 3,662 (I3662F) in the DNA-PKCS protein, and is strongly predicted by PolyPhen-2 to be damaging (score = 0.861).

Illustration of Mutations in
Gene & Protein
Protein Prediction

Figure 4. Domain structure of DNA-PKCS. The DNA-PKCS protein has three tetratricopeptide repeats (TPR) that are involved in protein-protein interactions such as with the Ku70/Ku80 heterodimer to form the DNA-PK complex.  The catalytic site is within the PIKK domain at the C-terminus.  FAT domains flank the PI3K/PI4K (PIKK) domain.  The N-terminus has three HEAT repeats. The daffy mutation is indicated. This image is interactive; click on each mutation to view more information.

The 12,674 base pair Prkdc transcript encodes the 4,128 amino acid, 471 kDa catalytic subunit of the DNA-PK complex, DNA-PKCS. DNA-PKCS is a serine/threonine kinase and member of the phosphatidylinositol 3-kinase-like kinases (PIKK) family (see worker) [reviewed in (1)]. DNA-PKCS has several domains that are essential for its function (Figure 4). A leucine rich region (LRR, aa 1501-1536) mediates the association of DNA-PKCS with C1D, a DNA-binding nuclear matrix-associated factor. The LRR facilitates the intrinsic binding of DNA-PKCS to DNA (2). The DNA-PKCS protein also contains three tetratricopeptide repeats (TPR) (aa 1720-1753, 2921-2954, 2956-2983) that are proposed to assist in protein-protein interactions.  A 380 amino acid region at the C terminus constitutes the catalytic domain, designated the PIKK domain, of DNA-PKCS (aa 3747-4015) (3). The PIKK domain is flanked by the FAT domain (named for its homology to FRAP, ATM and TRRAP, aa 2884-3539) and a FATC domain (FAT at the extreme C-terminus, aa 4096-4128) (3). The FAT and FATC domains occur in combination in all PIKK family members. The C-terminal region containing the FATC domain is essential for the kinase activity of DNA-PKCS (4-6). The N-terminal portion of the protein up to the FAT domain consists of HEAT (Huntingtin, Elongation factor 3, A subunit of protein phosphatase 2A and TOR1) repeats (amino acids 288-323, 1001-1037, and 1050-1086) (7). HEAT repeats are helical structural repeats that mediate protein-protein interactions (8).

The daffy mutation results in an isoleucine to phenylalanine substitution at position 3,662 (I3662F); I3662 is within an undefined domain between the FAT/FATC domain and the PIKK domain.

Please see the record clover for information about Prkdc.

Putative Mechanism

DNA-PKCS is the catalytic subunit of the DNA-PK complex and is essential for DNA double-strand break repair during nonhomologous end joining (NHEJ) and during the assembly of immune receptor genes (i.e., V(D)J recombination) in developing lymphocytes. Mutations in Prkdc are linked to severe combined immunodeficiency (SCID) in several animal models, a condition marked by lymphopenia, hypogammaglobulinemia, and impaired T and B cell-mediated functions (e.g. defective V(D)J recombination and reduced numbers of peripheral lymphocytes) (9-11). Mutations in Prkdc also exhibit uncapped telomeres and a large number of telomeric fusions, leading to genomic instability (9;12;13). In a spontaneous mouse model of SCID, a DNA-PKCS point mutation resulting in the loss of 83 C-terminal amino acids, a reduction in protein expression, and a block in lymphocyte development has been identified (14). The phenotype in the daffy mice indicate a loss of DNA-PKCS function. 

Primers PCR Primer
Daffy_pcr_F: AGAACTTGTAACTAGGAGCCATTC
Daffy_pcr_R: CCATGCTATTCCCCTTAAGAAGTG

Sequencing Primer
Daffy_seq_F: GGAGCCATTCTAATAATGACATGTC
Daffy_seq_R: GAAGTGTTCTTCTCACTATTGGATG
Genotyping

PCR program

1) 94°C 2:00
2) 94°C 0:30
3) 55°C 0:30
4) 72°C 1:00
5) repeat steps (2-4) 40x
6) 72°C 10:00
7) 4°C hold


The following sequence of 426 nucleotides is amplified (chromosome 16, + strand):


1   agaacttgta actaggagcc attctaataa tgacatgtct aaaggacgat aagacacaga
61  tgaaagagac attcgaaata ttttagaata ttgaaatatt cataagcttg gagctttttc
121 aaatgtataa aaggaatacg agtctttaca gtaaacttgt aatttccagg cgtttgggaa
181 agaatttgtc aagagctttg ggaatggtgg ttctaagctg cttacaatga aggtcgatga
241 cttttgtaag atcactggct cgctacttgt aaggatgaaa aaagactcaa agctacctgg
301 gaacctgaaa gagtattccc cctggatgag tgagttcaaa gcgcagttcc tgaaaaatga
361 actggagatt cctggtgagt taacatccaa tagtgagaag aacacttctt aaggggaata
421 gcatgg


Primer binding sites are underlined and the sequencing primers are highlighted; the mutated nucleotide is shown in red.

References
Science Writers Anne Murray
Illustrators Diantha La Vine
AuthorsXue Zhong, Jin Huk Choi, Zhao Zhang, and Bruce Beutler