Time-resolved analysis of DNA-protein interactions in living cells by UV laser pulses

被引：11

作者：

Nebbioso, Angela ^{[1
]}

Benedetti, Rosaria ^{[1
]}

Conte, Mariarosaria ^{[2
]}

Carafa, Vincenzo ^{[1
]}

De Bellis, Floriana ^{[1
,3
]}

Shaik, Jani ^{[3
]}

Matarese, Filomena ^{[3
]}

Della Ventura, Bartolomeo ^{[4
]}

Gesuele, Felice ^{[4
]}

Velotta, Raffaele ^{[4
]}

Martens, Joost H. A. ^{[1
,3
]}

Stunnenberg, Hendrik G. ^{[3
]}

Altucci, Carlo ^{[4
]}

Altucci, Lucia ^{[1
]}

机构：

[1] Univ Campania L Vanvitelli, Dipartimento Biochim Biofis & Patol Gen, Vico L De Crecchio 7, I-80138 Naples, IT, Italy

[2] IRCCS SDN, Via E Gianturco 113, I-80143 Naples, IT, Italy

[3] Radboud Univ Nijmegen, Dept Mol Biol, NCMLS, NL-6500 NL Nijmegen, Netherlands

[4] Univ Napoli Federico II, Dipartimento Fis, Via Cinthia, I-80100 Naples, IT, Italy

来源：

SCIENTIFIC REPORTS | 2017年 / 7卷

关键词：

ABSORBING AMINO-ACIDS; SOFT BIOLOGICAL TISSUES; CROSS-LINKING; THERMODYNAMIC RESPONSE; HDAC INHIBITORS; RECEPTOR; PHOTOCHEMISTRY; PHOTOPHYSICS; IDENTIFICATION; PATHWAYS;

D O I：

10.1038/s41598-017-12010-5

中图分类号：

O [数理科学和化学]; P [天文学、地球科学]; Q [生物科学]; N [自然科学总论];

学科分类号：

07 ; 0710 ; 09 ;

摘要：

Interactions between DNA and proteins are mainly studied through chemical procedures involving bi-functional reagents, mostly formaldehyde. Chromatin immunoprecipitation is used to identify the binding between transcription factors (TFs) and chromatin, and to evaluate the occurrence and impact of histone/DNA modifications. The current bottleneck in probing DNA-protein interactions using these approaches is caused by the fact that chemical crosslinkers do not discriminate direct and indirect bindings or short-lived chromatin occupancy. Here, we describe a novel application of UV laser-induced (L-) crosslinking and demonstrate that a combination of chemical and L-crosslinking is able to distinguish between direct and indirect DNA-protein interactions in a small number of living cells. The spatial and temporal dynamics of TF bindings to chromatin and their role in gene expression regulation may thus be assessed. The combination of chemical and L-crosslinking offers an exciting and unprecedented tool for biomedical applications.

引用

页数：13

共 36 条

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