RESUMO
DNA is traditionally seen as a linear sequence of instructions for cellular functions that are expressed through biochemical processes. Cellular DNA, however, is also organized as a complex hierarchical structure with a mosaic of mechanical features, and a growing body of evidence is now emerging to imply that these mechanical features are connected to genetic function. Mechanical tension, for instance, which must be felt by DNA within the heavily constrained and continually fluctuating cellular environment, can affect a number of regulatory processes implicating a role for biomechanics in gene expression complementary to that of biochemical regulation. In this article, we review evidence for such mechanical pathways of genetic regulation.
Assuntos
DNA/genética , Regulação da Expressão Gênica , Animais , Bioquímica/métodos , Fenômenos Biomecânicos , Biofísica/métodos , Cromatina/química , Cromossomos/ultraestrutura , Perfilação da Expressão Gênica , Humanos , Modelos Genéticos , Conformação de Ácido Nucleico , Transdução de Sinais , Transcrição GênicaRESUMO
Tethered particle motion (TPM) has become an important tool for single-molecule studies of biomolecules; however, concerns remain that the method may alter the dynamics of the biophysical process under study. We investigate the effect of the attached microsphere on an illustrative biological example: the formation and breakdown of protein-mediated DNA loops in the lac repressor system. By comparing data from a conventional TPM experiment with 800 nm polystyrene beads and dark-field TPM using 50 nm Au nanoparticles, we found that the lifetimes of the looped and unlooped states are only weakly modified, less than two-fold, by the presence of the large bead. This is consistent with our expectation of weak excluded-volume effects and hydrodynamic surface interactions from the cover glass and microsphere.
Assuntos
DNA/química , Proteínas/química , Fenômenos Biofísicos , Hidrodinâmica , Técnicas In Vitro , Cinética , Repressores Lac/química , Nanopartículas Metálicas , Microesferas , Modelos Moleculares , Movimento (Física) , Conformação de Ácido Nucleico , Tamanho da PartículaRESUMO
We describe a scanning-line optical tweezing technique with an asymmetric beam profile in the back focal plane of the microscope objective. The motion of a trapped particle along the scan line is studied as a function of beam asymmetry, and it is shown that this technique can be used to exert a constant lateral force on the particle, realizing purely optical constant-force tweezing. The observed effect is attributed in a geometric optics model to a non-zero lateral component of the scattering force.
Assuntos
Lasers , Microscopia/métodos , Óptica e Fotônica , Instrumentos Cirúrgicos , Fenômenos Biofísicos , Biofísica , Micromanipulação/métodos , Espalhamento de Radiação , Fatores de TempoRESUMO
We studied the thermal fluctuations of single DNA molecules with a novel optical tweezer based force spectroscopy technique. This technique combines femtonewton sensitivity with millisecond time resolution, surpassing the sensitivity of previous force measurements in aqueous solution with comparable bandwidth by a hundredfold. Our data resolve long-standing questions concerning internal hydrodynamics of the polymer and anisotropy in the molecular relaxation times and friction coefficients. The dynamics at high extension show interesting nonlinear behavior.