comparison of the btb superfamilies in physcomitrella
TRANSCRIPT
Comparison of the BTB Superfamilies in Physcomitrella patens and Arabidopsis thaliana
Jacob P. Janssen, Dr. Derek Gingerich,University of Wisconsin-Eau Claire
K
Target
BTBMotif
CUL3
RbxUb
C E2
BTB
The Arabidopsis and Physcomitrella BTB families encoded many similar BTB proteins
Figure 2. Grouping of the BTB subfamilies based on the nature of additional motifs outside of the BTB domain along with a protein diagram of representative members.
135987
2261782345479872811485710661652205
At5g64330
NPR1At3g48360
At5g19330At5g55000
NPH3
Ankyrin
TPRArmadilloPentapeptideOther
30At3g43700MATH 6
75321
26
17363
PhyscomitrellaArabidopsis
225
Other27
1 0 6 6 1 6 1 S DV T F L - V E GR R F Y AHR I AL - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - L AS S DAF R AMF D- - GG2 2 6 1 7 8 1 ADV K F E - V DGE L Y HAHK L V L - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - AAR S P V F K AQL F - - GP1 7 0 4 5 1 1 S DV V F E S L DGR K V Y GHR AV L - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - AS QS DV F K AMF S - - MP2 3 1 8 9 4 1 V DV E L QGV DGDR V HAHK AI L S V GWV L V DP S L C S E L GQV L L S T I HE S V R R R C P Y V I AE K K S V AS K S I E F E AMF R - - AD1 3 0 7 6 9 1 T DV T I C S AT G- C V GAHR AI L - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - AAR S P V F DS MF S - - HN1 6 5 6 6 0 1 ADV C F L - V GNE K F R C HR F V L G- - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - AR S E Y F K AR F S R T T G1 8 9 9 2 2 1 MDT S V V - L R S K T I HI S S AI L - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - AAK S P Y F Y K L F S - - NG9 8 7 2 8 1 ADL T V C T DDGS QI HV HS MV L - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - MAS S P V F K F Y L Q- - K E1 1 4 8 5 7 1 NDI V F H- L GGDAV P C NR E K I - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - AGL S MP F NT ML N- - GV1 3 5 9 8 7 1 S DI V V E - AGGMNF S L HK F P L - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - V AR S GR I R K L V ANL AD1 8 7 7 0 7 1 - S V V HL NV GGMMF AT T V DT L T - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - QR DT DS ML AV MF S GR HR1 0 4 5 4 5 1 - GR AK I E V GGR V F V T T MQT V - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - E K AGK DS HL Y K - - - T
1 0 6 6 1 6 K R L C E Y S MAQNL -2 2 6 1 7 8 R L L C E S K L C E NV -1 7 0 4 5 1 K S L C E QE I L QS L -2 3 1 8 9 4 L E V V C E DS L V K NM1 3 0 7 6 9 K E AC E S S L V DDI -1 6 5 6 6 0 K R AV T DAL L P QL -1 8 9 9 2 2 MR Y C S - R L L K NMP9 8 7 2 8 K QT C T V HF E QGL -1 1 4 8 5 7 K DAC DQNL AT F V -1 3 5 9 8 7 V AR T E AF L S E V V -1 8 7 7 0 7 V E NI T P F L C K K D-1 0 4 5 4 5 L R NAHGR AP L NG-
1 0 6 6 1 62 2 6 1 7 81 7 0 4 5 12 3 1 8 9 41 3 0 7 6 91 6 5 6 6 01 8 9 9 2 29 8 7 2 81 1 4 8 5 71 3 5 9 8 71 8 7 7 0 71 0 4 5 4 5
Y R E K E A- - - - - - - - - - - L DI E I P MR DR NS - - - - - - - - - - - DHI E I K MAE GT P - - - - - - - - - - - L HME MN T DR K V V - - - - - - - - - - - WNL E MP L ME K QS - - - - - - - - - - - AT V R I N F R E GV AGL AT NS GADS L L I L QE N MR E S E Q- - - - - - - - - R DV T L R I T V R K S R F P - - - - - - - - - - - AI DI F F L E AR MC D- - - - - - - - - I GF S K N S E NP NL - - - - - - - - - - - L QL T DV L HMDS K - - - - - - - - - - - E GAV F I V S QS GQ- - - - - - - - - - - - - - GWF
NI S WK V F E L MMR F I Y - - - - - - - - - - T GNV DMAT DNAQDL L R AADQY L L EDI E P P V F K AL L HF I Y R DS L P DT K E L I GAP S T S T L L AQHL L AAADR Y GL D- - - Y P AL DAF I F F F Y R - - - - - - - - AT V NQAV L R NHL V E L L QAADK Y GI E- - - Y V AL K AF V T F F Y T - - - - - - - - GR V S S S V L I E Y L T T L L DAAHK Y NV QDMC L E S C R AL L S F L Y G- - - - - - - - - NL K Y P DF R K HR V AL V R AAHK Y DI VDL S AS AF E K V L E Y I Y T - - - - - - - - - DS V K T V DL DE AE E L F DAAS R Y L L FQAE E S P L MDL L QF MY S - - - - - - - - - GR V QANT P AT V L DV L MAADK Y E V AGV P F HAV R C F L R Y L Y S - - - - - - - - S R C E R S DME E C AL HL V V L AHT Y MV PGI S V T GMR AV DHF S K T - - - - - - - - - GR L AR L S P E ML L E I L S F ANR F C C DP GGAE AF DL AAK F C Y G- - - - I NF E I T T S NV AV L R C AAE Y L DMT E S Y GE NDR DGT HF R HI L NWL R D- - - - - - - - - GV I P ML E I S AY QE L HR E AE Y Y QL MDR DP DMF NI L L NML R T G- - - - - - - V V Y P K P E S L GF L DR L I E E AT F Y GI E
GRAFDPTATNGE
LLL
LLCLLLLC
-
Sequence Alignment of Representatative Physcomitrella BTB Domains.
Introduction Following their synthesis, proteins in cells can be modified in a variety of ways. These modifications may modulate the protein’s activity, localization, interactions with other molecules, or stability. One such modification is the covalent attachment of a small globular protein called ubiquitin (Ub). Ubiquitination can change the activity or localization of a protein, if one Ub moiety is attached, or can lead to degradation of the protein if chains of Ubs are added. Selective degradation of key proteins by ubiquitination has been shown to be an important regulatory mechanism in many signaling pathways in eukaryotes. The attachment of Ub to target proteins is catalyzed by E3 Ub-ligases. One family of ligases are the BTB/Cullin 3 (CUL3) E3s. These complexes are composed of three proteins: CUL3, which acts as a backbone, RBX, which helps recruit the Ub, and a BTB (Bric-a-Brac/Tramtrack/Broad Complex) domain-containing protein (Fig. 1). The BTB motif allows the BTB protein to bind CUL3. Besides the BTB motif, BTB proteins contain other sequences (such as MATH or NPH3 domains) which recruit the target. BTB gene families are very diverse in eukaryotes. The sets of BTB proteins encoded in animal genomes are very different from those encoded in plants, for instance. We are interested in how the BTB gene family has changed during plant evolution. Dr. Gingerich has previously characterized the BTB family in a dicotyledenous flowering plant, Arabidopsis thaliana. In this studied, I have begun to characterize the family in a simple nonvascular moss, Physcomitrella patens. Significant differences in the BTB families might suggest that the two plants use the BTB/CUL3 E3s to target different proteins for degradation.
Conclusions
9 9
1 3 71 0 0
7999
1 0 2 1 0 2 1 0 2 1 0 2 1 0 8 1 0 4
79
BA1
D2RL
BPECJ
A2A2
39 known eukaryotic BTB domains used as initial queries
69 loci identified with < 0.0004 E-value
analysis and reannotation of gene sequences4 loci predicted to be nonfunctional
65 loci identified with < 0.0004 E-value
Figure 1 BTB/Cullin 3 E3 Ubiquitin-Ligases
77289
93266 61156
118125
130769
66017
97923
74560 124356150882
85405
87316
50789
42628 159693
219111
134565
30287
164153
100
100
100
100
93
100
6365
100
97
35
94
78
100
100
100
100
100
82
100
100
69100
10077
95
32
39
53
90
52
99
100
100
58
45
100
92
56
100
96
96
99
90
67
35
72 56
33
29 43
205 6
9
24 6
1
6
0
11
NPH3TAZ-CaM B DTPR
Armadillo
PentapeptideAP2Back 3
Physcomitrella ArabidopsisSubfamily size
626
3053
1
1380
Scop Domain 162
51
0
"A2" 43"D2" 21"F" 30"H" 56
Ankyrin 170451
106616
106476
165660
143618
226178
217835 98728
150321
223881
234547
231894 189922
122867
3260
121620 119190
31431
67811
231552
93641
41851
154574
21814
135987 127110
134520
12061 106949
12163
31592
2869 1322
22014
161287
114857
80508
50218
167201
111151
104545
148281
125607
187707 52205
52179
173
3
2
MATH
Other 5
00
Total 65
1
RPT2
NPH3, RPT3
PRL1-IFG
NPR1
AtPOB1
AtBT5AtBT4
AtBT3AtBT2AtBT1
At5g21010At3g43700
At3g03740At2g39760At5g19000
At3g06190At5g55000
At5g41330At3g09030At4g30940At2g24240
At5g19330At5g13060
At4g02680 EOL1At5g58550EOL2
At3g51770 ETO1At2g05330
At5g48510At4g04090
At2g40450At3g29740
At2g40440At3g56230
At1g01640At1g55760At1g21780
At4g08455
At4g37590 At2g23050At5g67440 At2g14820
At4g31820At5g47800
At1g67900At3g26490
At3g44820At5g10250
At3g50840At5g66560
At5g64330At1g30440
At5g03250 At5g13600At3g49970
At5g67385At5g48130
At2g30520At3g08570
At3g08660
At5g63160At3g48360
At1g05690At5g67480At4g37610
At2g30600At4g01160
At3g61600At2g46260
At5g45110At4g19660
At4g26120At1g64280
At3g57130At2g41370
At5g60050At1g63850At3g50780
At2g13690At3g05675
At5g48800At1g03010
At2g47860At3g22104
At1g50280At3g19850
At3g49900At3g03510
At5g17580At2g04740
At1g04390
10099
85
81
65
48
10026
4626
11100
9971
5213
933229
91
97
99
4646
37
42
6
100
9954
98
97
10096
69100
100
090
9760
58
2
5100
99
40
91100
6643
2799
2810
3
101997
3326
12
811
2851
100
79
7561
8586
8078
100
100
ARIA
BOP1BOP2
Phylogenetic Analysis of the BTB Superfamilies in Physcomitrella and Arabidopsis
Physcomitrella
Arabidopsis
Identification of BTB domain-containingproteins encoded by the Physcomitrella genome
BLAST search Physcomitrella patens genome v1.1 predicted protein sequences (from the DOE Joint Genome Institute Physcomitrella genome sequencing project)
Domain # of Genes Protein ID
TAZ zinc fingerETO1
# of Genes Protein ID
BA1
D2RL
BPECJ
A2A2
BA1
D2RL
BPECJ
A2A2
Figure 3. Sequence alignment of representative Physcomitrella BTB domains. The approximately 100 amino acid core BTB sequence from 12 representative Physcomitrella BTB proteins were aligned with ClustalW and displayed with Boxshade using a threshhold of 55%. Identical and conserved amino acids are shown with black and grey, respectively. Dashes denote gaps. Desginations to the left indicate the BTB family and the JGI number for each protein.
Some data presented in this poster was generated by Dr. Gingerich when he was funded by an NIH NRSA post-doctoral fellowship (#GM068361)
100
43
00
“K”“Q”
JCSTA2
Q I G H P R MN E A1
B L D2
K
J I H G F E D2
D1
C B A2
A1
Figure 4. Phylogenetic trees of the complete BTB protein superfamilies from Arabidopsis and Physcomitrella. The approximately 100 amino acid core BTB domain from all Arabidopsis and Physcomitrella BTB proteins were aligned byClustalW. The alignments were used to generate non-rooted phylogenetic trees with MEGA 4.0, using the Poisson distance method and a bootstrap value of 1000. The subfamilies identified from the phylogenetic analysis are marked on the bottom of the trees. Individual members of the tree are color-coded by the nature of the domain(s) either N- or C-terminal to the BTB domain. Where possible, designations for proteins previously identified by other methods are used.
-Physcomitrella patens encodes 65 putative BTB domain-containing proteins in its genome.
-There are a variety of different motifs/domains attached to BTB domains in the PhyscomitrellaBTB family, suggesting a number of different types of target proteins.
-The Physcomitrella and Arabidopsis BTB families are similar in overall size and share many similarBTB types, suggesting the two plants use the BTB family in similar ways.
-However, there are also several Physcomitrella and Arabidopsis-specific types, suggesting some specialization in each lineage.
-The BTB-NPH3 family in Arabidopsis is significantly larger than that in Physcomitrella (30 vs. 17), suggesting that there may have been an expansion of this subfamily as more complex plants evolved.