P20436 (RPAB3_YEAST) Reviewed, UniProtKB/Swiss-Prot

Names and origin

Protein names

Recommended name:
DNA-directed RNA polymerases I, II, and III subunit RPABC3
Short name=RNA polymerases I, II, and III subunit ABC3

Alternative name(s):
ABC14.4
ABC14.5
DNA-directed RNA polymerases I, II, and III 14.5 kDa polypeptide

Gene names

Name:	RPB8
Ordered Locus Names:	YOR224C
ORF Names:	YOR50-14

Organism

Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (Baker's yeast) [Reference proteome]

Taxonomic identifier

559292 [NCBI]

Taxonomic lineage

cellular organisms › Eukaryota › Opisthokonta › Fungi › Dikarya › Ascomycota › saccharomyceta › Saccharomycotina › Saccharomycetes › Saccharomycetales › Saccharomycetaceae › Saccharomyces › Saccharomyces cerevisiae

Protein attributes

Sequence length	146 AA.
Sequence status	Complete.
Protein existence	Evidence at protein level

General annotation (Comments)

Function	DNA-dependent RNA polymerase catalyzes the transcription of DNA into RNA using the four ribonucleoside triphosphates as substrates. Common component of RNA polymerases I, II and III which synthesize ribosomal RNA precursors, mRNA precursors and many functional non-coding RNAs, and small RNAs, such as 5S rRNA and tRNAs, respectively.
Subunit structure	Component of the RNA polymerase I (Pol I), RNA polymerase II (Pol II) and RNA polymerase III (Pol III) complexes consisting of 14, 12 and 17 subunits, respectively. Directly interacts with POLR2A. Ref.7
Subcellular location	Nucleus Ref.8.
Miscellaneous	Present with 6210 molecules/cell in log phase SD medium.
Sequence similarities	Belongs to the eukaryotic RPB8 RNA polymerase subunit family.

Ontologies

Keywords
Biological process	Transcription
Cellular component	DNA-directed RNA polymerase Nucleus
Ligand	DNA-binding
PTM	Phosphoprotein
Technical term	3D-structure Complete proteome Reference proteome
Gene Ontology (GO)
Biological_process	tRNA transcription from RNA polymerase III promoter Inferred from direct assay PubMed 3905793. Source: SGD transcription from RNA polymerase I promoter Inferred from physical interaction PubMed 11486042. Source: SGD transcription from RNA polymerase II promoter Inferred from physical interaction PubMed 11486042. Source: SGD
Cellular_component	DNA-directed RNA polymerase I complex Inferred from direct assay PubMed 11486042PubMed 9837969. Source: SGD DNA-directed RNA polymerase II, core complex Inferred from direct assay PubMed 1331084PubMed 2183013Ref.1. Source: SGD DNA-directed RNA polymerase III complex Inferred from direct assay PubMed 10611227PubMed 3905793. Source: SGD
Molecular_function	DNA binding Inferred from electronic annotation. Source: UniProtKB-KW DNA-directed RNA polymerase activity Inferred from electronic annotation. Source: UniProtKB-KW
Complete GO annotation...

Binary interactions

With	Entry	#Exp.	IntAct	Notes
RSP5	P39940	3	EBI-15794,EBI-16219

Sequence annotation (Features)

Feature key

Position(s)

Length

Description

Graphical view

Feature identifier

Molecule processing

Chain

1 – 146

146

DNA-directed RNA polymerases I, II, and III subunit RPABC3

PRO_0000074001

Regions

Region

16 – 39

24

Non-specific ssDNA binding By similarity

Amino acid modifications

Modified residue

68

1

Phosphothreonine Ref.10

Secondary structure

1

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146

Helix Strand Turn

Details...

Beta strand

7 – 16

10

Beta strand

21 – 32

12

Beta strand

37 – 43

7

Helix

44 – 46

3

Beta strand

54 – 58

5

Turn

84 – 86

3

Beta strand

93 – 101

9

Beta strand

108 – 110

3

Beta strand

112 – 118

7

Beta strand

121 – 128

8

Turn

129 – 133

5

Beta strand

136 – 138

3

Beta strand

140 – 145

6

Sequences

	Sequence		Length	Mass (Da)
	P20436 [UniParc]. Last modified February 1, 1991. Version 1. Checksum: 5B71693FE6FD8863	FASTA	146	16,511
	10 20 30 40 50 60 MSNTLFDDIF QVSEVDPGRY NKVCRIEAAS TTQDQCKLTL DINVELFPVA AQDSLTVTIA 70 80 90 100 110 120 SSLNLEDTPA NDSSATRSWR PPQAGDRSLA DDYDYVMYGT AYKFEEVSKD LIAVYYSFGG 130 140 LLMRLEGNYR NLNNLKQENA YLLIRR

References

	« Hide 'large scale' referencesShow 'large scale' references »
[1]	"Subunits shared by eukaryotic nuclear RNA polymerases." Woychik N.A., Liao S.-M., Kolodziej P.A., Young R.A. Genes Dev. 4:313-323(1990) [PubMed] [Europe PMC] [Abstract] Cited for: NUCLEOTIDE SEQUENCE [GENOMIC DNA].
[2]	"Sequence and analysis of a 33 kb fragment from the right arm of chromosome XV of the yeast Saccharomyces cerevisiae." Galisson F., Dujon B. Yeast 12:877-885(1996) [PubMed] [Europe PMC] [Abstract] Cited for: NUCLEOTIDE SEQUENCE [GENOMIC DNA]. Strain: ATCC 96604 / S288c / FY1679.
[3]	"The nucleotide sequence of Saccharomyces cerevisiae chromosome XV." Dujon B., Albermann K., Aldea M., Alexandraki D., Ansorge W., Arino J., Benes V., Bohn C., Bolotin-Fukuhara M., Bordonne R., Boyer J., Camasses A., Casamayor A., Casas C., Cheret G., Cziepluch C., Daignan-Fornier B., Dang V.-D. , de Haan M., Delius H., Durand P., Fairhead C., Feldmann H., Gaillon L., Galisson F., Gamo F.-J., Gancedo C., Goffeau A., Goulding S.E., Grivell L.A., Habbig B., Hand N.J., Hani J., Hattenhorst U., Hebling U., Hernando Y., Herrero E., Heumann K., Hiesel R., Hilger F., Hofmann B., Hollenberg C.P., Hughes B., Jauniaux J.-C., Kalogeropoulos A., Katsoulou C., Kordes E., Lafuente M.J., Landt O., Louis E.J., Maarse A.C., Madania A., Mannhaupt G., Marck C., Martin R.P., Mewes H.-W., Michaux G., Paces V., Parle-McDermott A.G., Pearson B.M., Perrin A., Pettersson B., Poch O., Pohl T.M., Poirey R., Portetelle D., Pujol A., Purnelle B., Ramezani Rad M., Rechmann S., Schwager C., Schweizer M., Sor F., Sterky F., Tarassov I.A., Teodoru C., Tettelin H., Thierry A., Tobiasch E., Tzermia M., Uhlen M., Unseld M., Valens M., Vandenbol M., Vetter I., Vlcek C., Voet M., Volckaert G., Voss H., Wambutt R., Wedler H., Wiemann S., Winsor B., Wolfe K.H., Zollner A., Zumstein E., Kleine K. Nature 387:98-102(1997) [PubMed] [Europe PMC] [Abstract] Cited for: NUCLEOTIDE SEQUENCE [LARGE SCALE GENOMIC DNA]. Strain: ATCC 96604 / S288c / FY1679.
[4]	Saccharomyces Genome Database Submitted (DEC-2009) to the EMBL/GenBank/DDBJ databases Cited for: GENOME REANNOTATION. Strain: ATCC 204508 / S288c.
[5]	"Approaching a complete repository of sequence-verified protein-encoding clones for Saccharomyces cerevisiae." Hu Y., Rolfs A., Bhullar B., Murthy T.V.S., Zhu C., Berger M.F., Camargo A.A., Kelley F., McCarron S., Jepson D., Richardson A., Raphael J., Moreira D., Taycher E., Zuo D., Mohr S., Kane M.F., Williamson J. , Simpson A.J.G., Bulyk M.L., Harlow E., Marsischky G., Kolodner R.D., LaBaer J. Genome Res. 17:536-543(2007) [PubMed] [Europe PMC] [Abstract] Cited for: NUCLEOTIDE SEQUENCE [GENOMIC DNA]. Strain: ATCC 204508 / S288c.
[6]	"The yeast RNA polymerase III transcription machinery: a paradigm for eukaryotic gene activation." Chedin S., Ferri M.L., Peyroche G., Andrau J.-C., Jourdain S., Lefebvre O., Werner M., Carles C., Sentenac A. Cold Spring Harb. Symp. Quant. Biol. 63:381-389(1998) [PubMed] [Europe PMC] [Abstract] Cited for: REVIEW ON THE RNA POL III COMPLEX.
[7]	"The A14-A43 heterodimer subunit in yeast RNA pol I and their relationship to Rpb4-Rpb7 pol II subunits." Peyroche G., Levillain E., Siaut M., Callebaut I., Schultz P., Sentenac A., Riva M., Carles C. Proc. Natl. Acad. Sci. U.S.A. 99:14670-14675(2002) [PubMed] [Europe PMC] [Abstract] Cited for: IDENTIFICATION IN THE RNA POL I COMPLEX.
[8]	"Global analysis of protein localization in budding yeast." Huh W.-K., Falvo J.V., Gerke L.C., Carroll A.S., Howson R.W., Weissman J.S., O'Shea E.K. Nature 425:686-691(2003) [PubMed] [Europe PMC] [Abstract] Cited for: SUBCELLULAR LOCATION [LARGE SCALE ANALYSIS].
[9]	"Global analysis of protein expression in yeast." Ghaemmaghami S., Huh W.-K., Bower K., Howson R.W., Belle A., Dephoure N., O'Shea E.K., Weissman J.S. Nature 425:737-741(2003) [PubMed] [Europe PMC] [Abstract] Cited for: LEVEL OF PROTEIN EXPRESSION [LARGE SCALE ANALYSIS].
[10]	"A multidimensional chromatography technology for in-depth phosphoproteome analysis." Albuquerque C.P., Smolka M.B., Payne S.H., Bafna V., Eng J., Zhou H. Mol. Cell. Proteomics 7:1389-1396(2008) [PubMed] [Europe PMC] [Abstract] Cited for: PHOSPHORYLATION [LARGE SCALE ANALYSIS] AT THR-68, MASS SPECTROMETRY.
[11]	"Eukaryotic RNA polymerase subunit RPB8 is a new relative of the OB family." Krapp S., Kelly G., Reischi J., Weinzierl R.O.J., Matthews S. Nat. Struct. Biol. 5:110-114(1998) [PubMed] [Europe PMC] [Abstract] Cited for: STRUCTURE BY NMR.
[12]	"RNA polymerase II/TFIIF structure and conserved organization of the initiation complex." Chung W.H., Craighead J.L., Chang W.H., Ezeokonkwo C., Bareket-Samish A., Kornberg R.D., Asturias F.J. Mol. Cell 12:1003-1013(2003) [PubMed] [Europe PMC] [Abstract] Cited for: ELECTRON MICROSCOPY OF THE RNA POL II/TFIIF COMPLEX.
[13]	"Structural basis of transcription: RNA polymerase II at 2.8 A resolution." Cramer P., Bushnell D.A., Kornberg R.D. Science 292:1863-1876(2001) [PubMed] [Europe PMC] [Abstract] Cited for: X-RAY CRYSTALLOGRAPHY (2.8 ANGSTROMS) OF THE RNA POL II CORE COMPLEX.
[14]	"Structural basis of transcription: an RNA polymerase II elongation complex at 3.3 A resolution." Gnatt A.L., Cramer P., Fu J., Bushnell D.A., Kornberg R.D. Science 292:1876-1882(2001) [PubMed] [Europe PMC] [Abstract] Cited for: X-RAY CRYSTALLOGRAPHY (3.3 ANGSTROMS) OF THE RNA POL II CORE COMPLEX.
[15]	"Structural basis of transcription: alpha-amanitin-RNA polymerase II cocrystal at 2.8 A resolution." Bushnell D.A., Cramer P., Kornberg R.D. Proc. Natl. Acad. Sci. U.S.A. 99:1218-1222(2002) [PubMed] [Europe PMC] [Abstract] Cited for: X-RAY CRYSTALLOGRAPHY (2.8 ANGSTROMS) OF THE RNA POL II CORE COMPLEX IN COMPLEX WITH ALPHA-AMANITIN.
[16]	"Architecture of the RNA polymerase II-TFIIS complex and implications for mRNA cleavage." Kettenberger H., Armache K.J., Cramer P. Cell 114:347-357(2003) [PubMed] [Europe PMC] [Abstract] Cited for: X-RAY CRYSTALLOGRAPHY (3.8 ANGSTROMS) OF THE RNA POL II COMPLEX IN COMPLEX WITH DST1.
[17]	"Architecture of initiation-competent 12-subunit RNA polymerase II." Armache K.J., Kettenberger H., Cramer P. Proc. Natl. Acad. Sci. U.S.A. 100:6964-6968(2003) [PubMed] [Europe PMC] [Abstract] Cited for: X-RAY CRYSTALLOGRAPHY (4.2 ANGSTROMS) OF THE RNA POL II COMPLEX.
[18]	"Complete, 12-subunit RNA polymerase II at 4.1-A resolution: implications for the initiation of transcription." Bushnell D.A., Kornberg R.D. Proc. Natl. Acad. Sci. U.S.A. 100:6969-6973(2003) [PubMed] [Europe PMC] [Abstract] Cited for: X-RAY CRYSTALLOGRAPHY (4.1 ANGSTROMS) OF THE RNA POL II CORE COMPLEX.
[19]	"Structural basis of transcription: nucleotide selection by rotation in the RNA polymerase II active center." Westover K.D., Bushnell D.A., Kornberg R.D. Cell 119:481-489(2004) [PubMed] [Europe PMC] [Abstract] Cited for: X-RAY CRYSTALLOGRAPHY (2.3 ANGSTROMS) OF THE RNA POL II CORE COMPLEX.
[20]	"Complete RNA polymerase II elongation complex structure and its interactions with NTP and TFIIS." Kettenberger H., Armache K.J., Cramer P. Mol. Cell 16:955-965(2004) [PubMed] [Europe PMC] [Abstract] Cited for: X-RAY CRYSTALLOGRAPHY (4.5 ANGSTROMS).
[21]	"Structural basis of transcription: an RNA polymerase II-TFIIB cocrystal at 4.5 Angstroms." Bushnell D.A., Westover K.D., Davis R.E., Kornberg R.D. Science 303:983-988(2004) [PubMed] [Europe PMC] [Abstract] Cited for: X-RAY CRYSTALLOGRAPHY (4.5 ANGSTROMS) OF THE RNA POL II CORE COMPLEX.
[22]	"Structures of complete RNA polymerase II and its subcomplex, Rpb4/7." Armache K.J., Mitterweger S., Meinhart A., Cramer P. J. Biol. Chem. 280:7131-7134(2005) [PubMed] [Europe PMC] [Abstract] Cited for: X-RAY CRYSTALLOGRAPHY (3.8 ANGSTROMS) OF THE RNA POL II COMPLEX.
[23]	"Structural biology of RNA polymerase III: subcomplex C17/25 X-ray structure and 11 subunit enzyme model." Jasiak A.J., Armache K.J., Martens B., Jansen R.P., Cramer P. Mol. Cell 23:71-81(2006) [PubMed] [Europe PMC] [Abstract] Cited for: 3D-STRUCTURE MODELING OF THE POL III CORE COMPLEX.
[24]	"Structure of an RNA polymerase II-RNA inhibitor complex elucidates transcription regulation by noncoding RNAs." Kettenberger H., Eisenfuhr A., Brueckner F., Theis M., Famulok M., Cramer P. Nat. Struct. Mol. Biol. 13:44-48(2006) [PubMed] [Europe PMC] [Abstract] Cited for: X-RAY CRYSTALLOGRAPHY (3.8 ANGSTROMS) OF THE RNA POL II COMPLEX IN COMPLEX WITH INHIBITING NON-CODING RNA.
[25]	"Phasing RNA polymerase II using intrinsically bound Zn atoms: an updated structural model." Meyer P.A., Ye P., Zhang M., Suh M.H., Fu J. Structure 14:973-982(2006) [PubMed] [Europe PMC] [Abstract] Cited for: X-RAY CRYSTALLOGRAPHY (4.15 ANGSTROMS) OF THE RNA POL II COMPLEX.
+	Additional computationally mapped references.

Cross-references

Sequence databases

EMBL
GenBank
DDBJ

X53289 Genomic DNA. Translation: CAA37383.1.
X92441 Genomic DNA. Translation: CAA63187.1.
Z75132 Genomic DNA. Translation: CAA99443.1.
AY693087 Genomic DNA. Translation: AAT93106.1.
BK006948 Genomic DNA. Translation: DAA10995.1.

PIR

C34588.

RefSeq

NP_014867.1. NM_001183643.1.

3D structure databases

PDBe
RCSB PDB
PDBj

Entry	Method	Resolution (Å)	Chain	Positions	PDBsum
1A1D	NMR	-	A	1-146	[»]
1I3Q	X-ray	3.10	H	1-146	[»]
1I50	X-ray	2.80	H	1-146	[»]
1I6H	X-ray	3.30	H	1-146	[»]
1K83	X-ray	2.80	H	1-146	[»]
1NIK	X-ray	4.10	H	1-146	[»]
1NT9	X-ray	4.20	H	1-146	[»]
1PQV	X-ray	3.80	H	1-146	[»]
1R5U	X-ray	4.50	H	1-146	[»]
1R9S	X-ray	4.25	H	1-146	[»]
1R9T	X-ray	3.50	H	1-146	[»]
1SFO	X-ray	3.61	H	1-146	[»]
1TWA	X-ray	3.20	H	1-146	[»]
1TWC	X-ray	3.00	H	1-146	[»]
1TWF	X-ray	2.30	H	1-146	[»]
1TWG	X-ray	3.30	H	1-146	[»]
1TWH	X-ray	3.40	H	1-146	[»]
1WCM	X-ray	3.80	H	1-146	[»]
1Y1V	X-ray	3.80	H	1-146	[»]
1Y1W	X-ray	4.00	H	1-146	[»]
1Y1Y	X-ray	4.00	H	1-146	[»]
1Y77	X-ray	4.50	H	1-146	[»]
2B63	X-ray	3.80	H	1-146	[»]
2B8K	X-ray	4.15	H	1-146	[»]
2E2H	X-ray	3.95	H	1-146	[»]
2E2I	X-ray	3.41	H	1-146	[»]
2E2J	X-ray	3.50	H	1-146	[»]
2JA5	X-ray	3.80	H	1-146	[»]
2JA6	X-ray	4.00	H	1-146	[»]
2JA7	X-ray	3.80	H/T	1-146	[»]
2JA8	X-ray	3.80	H	1-146	[»]
2NVQ	X-ray	2.90	H	1-146	[»]
2NVT	X-ray	3.36	H	1-146	[»]
2NVX	X-ray	3.60	H	1-146	[»]
2NVY	X-ray	3.40	H	1-146	[»]
2NVZ	X-ray	4.30	H	1-146	[»]
2R7Z	X-ray	3.80	H	1-146	[»]
2R92	X-ray	3.80	H	1-146	[»]
2R93	X-ray	4.00	H	1-146	[»]
2VUM	X-ray	3.40	H	1-146	[»]
2YU9	X-ray	3.40	H	1-146	[»]
3CQZ	X-ray	2.80	H	1-146	[»]
3FKI	X-ray	3.88	H	1-146	[»]
3GTG	X-ray	3.78	H	1-146	[»]
3GTJ	X-ray	3.42	H	1-146	[»]
3GTK	X-ray	3.80	H	1-146	[»]
3GTL	X-ray	3.38	H	1-146	[»]
3GTM	X-ray	3.80	H	1-146	[»]
3GTO	X-ray	4.00	H	1-146	[»]
3GTP	X-ray	3.90	H	1-146	[»]
3GTQ	X-ray	3.80	H	1-146	[»]
3H3V	X-ray	4.00	I	1-146	[»]
3HOU	X-ray	3.20	H/T	1-146	[»]
3HOV	X-ray	3.50	H	1-146	[»]
3HOW	X-ray	3.60	H	1-146	[»]
3HOX	X-ray	3.65	H	1-146	[»]
3HOY	X-ray	3.40	H	1-146	[»]
3HOZ	X-ray	3.65	H	1-146	[»]
3I4M	X-ray	3.70	H	1-146	[»]
3I4N	X-ray	3.90	H	1-146	[»]
3J1N	electron microscopy	16.00	H	1-146	[»]
3K1F	X-ray	4.30	H	1-146	[»]
3K7A	X-ray	3.80	H	1-146	[»]
3M3Y	X-ray	3.18	H	1-146	[»]
3M4O	X-ray	3.57	H	1-146	[»]
3PO2	X-ray	3.30	H	1-146	[»]
3PO3	X-ray	3.30	H	1-146	[»]
3QT1	X-ray	4.30	H	1-146	[»]
3RZD	X-ray	3.30	H	1-146	[»]
3RZO	X-ray	3.00	H	1-146	[»]
3S14	X-ray	2.85	H	1-146	[»]
3S15	X-ray	3.30	H	1-146	[»]
3S16	X-ray	3.24	H	1-146	[»]
3S17	X-ray	3.20	H	1-146	[»]
3S1M	X-ray	3.13	H	1-146	[»]
3S1N	X-ray	3.10	H	1-146	[»]
3S1Q	X-ray	3.30	H	1-146	[»]
3S1R	X-ray	3.20	H	1-146	[»]
3S2D	X-ray	3.20	H	1-146	[»]
3S2H	X-ray	3.30	H	1-146	[»]
4A3B	X-ray	3.50	H	1-146	[»]
4A3C	X-ray	3.50	H	1-146	[»]
4A3D	X-ray	3.40	H	1-146	[»]
4A3E	X-ray	3.40	H	1-146	[»]
4A3F	X-ray	3.50	H	1-146	[»]
4A3G	X-ray	3.50	H	1-146	[»]
4A3I	X-ray	3.80	H	1-146	[»]
4A3J	X-ray	3.70	H	1-146	[»]
4A3K	X-ray	3.50	H	1-146	[»]
4A3L	X-ray	3.50	H	1-146	[»]
4A3M	X-ray	3.90	H	1-146	[»]
4A93	X-ray	3.40	H	1-146	[»]
4BBR	X-ray	3.40	H	1-146	[»]
4BBS	X-ray	3.60	H	1-146	[»]

ProteinModelPortal

P20436.

SMR

P20436. Positions 1-146.

ModBase

Search...

Protein-protein interaction databases

DIP

DIP-2195N.

IntAct

P20436. 19 interactions.

MINT

MINT-507506.

STRING

4932.YOR224C.

Proteomic databases

PaxDb

P20436.

PeptideAtlas

P20436.

Protocols and materials databases

StructuralBiologyKnowledgebase

Search...

Genome annotation databases

EnsemblFungi

YOR224C; YOR224C; YOR224C.

GeneID

854399.

KEGG

sce:YOR224C.

Organism-specific databases

CYGD

YOR224c.

SGD

S000005750. RPB8.

Phylogenomic databases

eggNOG

NOG279425.

GeneTree

ENSGT00390000018195.

HOGENOM

HOG000175572.

KO

K03016.

OMA

SFKMELI.

OrthoDB

EOG4QJVXT.

Enzyme and pathway databases

Reactome

REACT_83470. Gene Expression.
REACT_87991. Transcription.

Gene expression databases

Genevestigator

P20436.

GermOnline

YOR224C. Saccharomyces cerevisiae.

Family and domain databases

Gene3D

2.40.50.140. 1 hit.

InterPro

IPR012340. NA-bd_OB-fold.
IPR005570. RNA_pol_Rpb8.
[Graphical view]

PANTHER

PTHR10917. PTHR10917. 1 hit.

Pfam

PF03870. RNA_pol_Rpb8. 1 hit.
[Graphical view]

PIRSF

PIRSF000779. RNA_pol_Rpb8. 1 hit.

SMART

SM00658. RPOL8c. 1 hit.
[Graphical view]

SUPFAM

SSF50249. Nucleic_acid_OB. 1 hit.

ProtoNet

Search...

Other

EvolutionaryTrace

P20436.

NextBio

976571.

Entry information

Entry name

RPAB3_YEAST

Accession

Primary (citable) accession number: P20436
Secondary accession number(s): D6W2S9

Entry history

Integrated into UniProtKB/Swiss-Prot:	February 1, 1991
Last sequence update:	February 1, 1991
Last modified:	May 1, 2013
This is version 136 of the entry and version 1 of the sequence. [Complete history]

Entry status

Reviewed (UniProtKB/Swiss-Prot)

Annotation program

Fungal Protein Annotation Program