El papel de la curvatura de DNA en la regulación

El papel de la curvatura de DNA en la regulación

Clues and consequences of DNA bending in transcription

• Nature of DNA bending. Bendability vs curved DNA

• Clues of DNA bending

– Inducer/ inhibitor of protein-DNA interactions

– Catalyst of protein-protein interactions

– DNA chaperones

• Consequences of DNA bending

– Channeling signals through promoter architecture

– Helping response-amplification signals

– Avoiding transcriptional promiscuity

• DNA bending: A new signal-transduction mechanism?

Curved DNA

Wedge model Structural discontinuitiesmodel

Bendability

• Bendability: the ability of specific short

sequences to assume preferentially

conformations that accommodate the

deformation associated with protein-

induced bending.

Bendability vs curved DNA

• Curved DNA is deformed even in the absence of

external forces, thereby resulting in a very rigid

structure.

• Bendable DNA allows a mixture of many different

conformational states, the equilibrium of which can

be displaced toward one specific form by external

forces such as proteins interacting with them.

Protein-mediated DNA bending

• Neutralization of charges in

the DNA backbone

• Setting up extended

protein-DNA contacts

• Intercalation of protein side

chains in the minor groove

Bend DNA is important for DNA-protein interactions

• CAP: Correlation between bendability of CAP site

and the affinity of the protein (Gartenberg and

Crothers, 1988)

• Bacterial 70-RNA polymerase induces a strong

bend in the promoter upon binding (Pérez-Martín

and Espinosa, 1994)

• TBP and holo-TFIID bend DNA (Starr et al., 1995)

Clues of DNA bending in transcription

• Inducer/ inhibitor of protein-DNA interactions

• Catalyst of protein-protein interactions

• DNA chaperones

DNA bending as an inducer or inhibitor of DNA-protein

interactions• Structural synergy:

– Pre-curved CAP DNA binding sites

– HMG1 and the human progesterone receptor

• Structural inhibition:

– Out of phase RepA-induced bends

Structural synergy at the CAP-binding sites (Kahn and Crothers, 1992)

Structural synergy between HMG1 and PR (Oñate et al., 1994)

HMG1

PR

Structural inhibition (Pérez-Martín and Espinosa, 1991)

RNApolRNApol

RNApolRNApol+ RepA




• DNA chaperones

DNA bending as a catalyst of protein-protein interactions

• Long distances: – IHF

– histones

• Short distances:– bendability: XylR

enhancer– extra factors: LEF-

1 enhancer

Distance (bp)

|10

|100

|1000

Free e

nerg

y

XylR enhancer (Pérez-Martín and de Lorenzo, 1996)

30 bp

ATP

Bendable DNA

LEF-1 enhancer in TCR (Giese et al., 1992)

LEF-1

PEBP2

Ets-1

ATF/CREB

DNA bending as a catalyst of protein-protein interactions

• Long distances: – IHF

– histones

• Short distances:– bendability: XylR

enhancer– extra factors: LEF-

1 enhancer

Distance (bp)

|10

|100

|1000

Free e

nerg

y

IHF at Pu promoter (Pérez-Martín and de Lorenzo, 1996)

XylR54-RNAP

54-RNAPIHF

>200 bp

Nucleosome positioning at the Drosophila hsp26 promoter (Thomas

and Elgin, 1988)

TATA boxHSTF-box GAGA-boxGAGA-box HSTF-box

-100-200-300-400

RNApolIInucleosome




• DNA chaperones

DNA chaperones (Travers, 1994)

• DNA chaperones are DNA-bending proteins that stabilize an otherwise loose structure in a particular conformation which sustains the assembly of additional proteins into a higher-order complex, being displaced away from the DNA in the final assembly.

DNA chaperones: HMG1 and PR (Oñate et al., 1994)

PR HMG1

DNA chaperones: HU at Ps promoter (Pérez-Martín and de Lorenzo, 1995)

54-RNAP

XylR

54-RNAPHU

54-RNAP

Consequences of DNA bending in transcription

• Channeling signals through promoter architecture

• Helping response-amplification signals

• Avoiding transcriptional promiscuity

Channeling signals through promoter architecture

• Co-activation

– CAP and MalT

– nucleosome in Xenopus vitellogenin B1 promoter

• Anti-repression

– CAP in ParaBAD

• Anti-induction

– YY1 in c-fos promoter

– IHF in nac promoter

cAMP

Co-activation: MalT and CAP at PmalE-PmalK in E.coli (Richet et al., 1991)

PmalE

PmalK

RNApol

RNApol

MalTCAP

Maltose

Co-activation: Xenopus vitellogenin B1 promoter (Schild et al., 1993)

Oestrogenreceptor

RNApolIIHNF3

NF1Hormone

Cell-type

Anti-repression: AraC and CAP at ParaBAD in E.coli (Lobell and Schleif,

1991)

RNApol

RNApolRNApol

AraC

CAPcAMP Arabinose

Anti-induction: YY1 at the c- fos promoter (Natesan and Gilman, 1993)

CREBP

YY1

RNApolII

cAMP

Cell status

Anti-induction: Nac at the nac promoter from K. aerogenes (Feng et al., 1995)

PnacRNApol

HU

Nac

NtrC

Nitrogenstatus Nac levels





DNA bending in response-amplification

mechanisms

Stimulation of lysis/lisogeny of phage Mu by DNA bending (Goosen, van de Putte, 1995)

O1 IHF O2 O3

Pe

RNApol Pe

O1

O2 O3

LYSIS LYSOGENY





DNA bending and transcriptional promiscuity

• Mechanisms to suppress non-specific activation at enhancers: restrictors

? ?

Restrictor: a new role of IHF in Pu promoter (Pérez-Martín and de Lorenzo,

1995)

XylR

IHF54-RNAP

IHF54-RNAP

+IHF

54-RNAP

-IHF

Clues and consequences of DNA bending in transcription

DNA bending: A new signal-transduction mechanism?

Characteristics of signal-transduction mechanisms

• Integration of signals

• Amplification of signals

• Specificity of signals

Integration of signals

Kinase3

Kinase2

Kinase1SignalA

SignalB

SignalC





Amplification of signals

Kinase2

Kinase1

Kinase2

Kinase3Kinase3 Kinase3Kinase3





Specificity of signals(Scaffolding)

Kinase3

Kinase2

Kinase1

SignalA SignalB

Kinase3

Kinase2

Kinase1

Designing promoters “ a la carte”

Integration

AmplificationSpecificity

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El papel de la curvatura de DNA en la regulación