Original gels illustrated in Figures 5, 6 and S5 illustrating hydroxyl-radical footprinting and cross-linking results.
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Research Interests: Computational Biology, Biology, Recognition, Medicine, Biological Sciences, and 14 moreSaccharomyces cerevisiae, Environmental Sciences, Humans, Diseases, NF-kappa B, Genome, Single Nucleotide Polymorphism, Quantitative Trait Loci, Sites, Base Sequence, In Vitro Selection, alleles, binding sites, and Specificities
Research Interests: Microscopy, Protein Folding, Biology, Atomic Force Microscopy, Medicine, and 15 moreBiological Sciences, DNA, Mutation, Animals, Nucleosomes, Chromatin, Histones, Nucleosome, Genetic variation, Amino Acid Sequence, Protein Quaternary Structure, Molecular Sequence Data, binding sites, Medical and Health Sciences, and In Vitro Techniques
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In this study, we examined the effect of acetylation of the NH 2 tails of core histones on their binding to nucleosomal DNA in the absence or presence of bound transcription factors. To do this, we used a novel UV laser-induced... more
In this study, we examined the effect of acetylation of the NH 2 tails of core histones on their binding to nucleosomal DNA in the absence or presence of bound transcription factors. To do this, we used a novel UV laser-induced protein-DNA cross-linking technique, combined with immunochemical and molecular biology approaches. Nucleosomes containing one or five GAL4 binding sites were reconstituted with hypoacetylated or hyperacetylated core histones. Within these reconstituted particles, UV laser-induced histone-DNA cross-linking was found to occur only via the nonstructured histone tails and thus presented a unique tool for studying histone tail interactions with nucleosomal DNA. Importantly, these studies demonstrated that the NH 2 tails were not released from nucleosomal DNA upon histone acetylation, although some weakening of their interactions was observed at elevated ionic strengths. Moreover, the binding of up to five GAL4-AH dimers to nucleosomes occupying the central 90 bp ...
Research Interests: Cellular Biology, Biology, Medicine, Biological Sciences, DNA, and 15 moreMolecular and cellular biology, Lasers, Animals, Histone Methylation, Histones, Chickens, Acetylation, Histone Code, Histone, Ionic Strength, Erythrocytes, DNA binding proteins, Binding Site, Histone Acetylation, and Medical and Health Sciences
ABSTRACT
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Linker histones associate with nucleosomes to promote the formation of higher-order chromatin structure, but the underlying molecular details are unclear. We investigated the structure of a 197 bp nucleosome bearing symmetric 25 bp linker... more
Linker histones associate with nucleosomes to promote the formation of higher-order chromatin structure, but the underlying molecular details are unclear. We investigated the structure of a 197 bp nucleosome bearing symmetric 25 bp linker DNA arms in complex with vertebrate linker histone H1. We determined electron cryo-microscopy (cryo-EM) and crystal structures of unbound and H1-bound nucleosomes and validated these structures by site-directed protein cross-linking and hydroxyl radical footprinting experiments. Histone H1 shifts the conformational landscape of the nucleosome by drawing the two linkers together and reducing their flexibility. The H1 C-terminal domain (CTD) localizes primarily to a single linker, while the H1 globular domain contacts the nucleosome dyad and both linkers, associating more closely with the CTD-distal linker. These findings reveal that H1 imparts a strong degree of asymmetry to the nucleosome, which is likely to influence the assembly and architecture ...
Research Interests: Biological Sciences, DNA, Humans, Animals, Nucleosomes, and 13 moreChromatin, Chromatin structure, Histones, Nucleosome, X ray diffraction, Time Factors, Xenopus laevis, Structure activity Relationship, Protein Binding, binding sites, Linker histone, chromatosome structure, and Medical and Health Sciences
RNA polymerase V (Pol V) long noncoding RNAs (lncRNAs) have been proposed to guide ARGONAUTE4 (AGO4) to chromatin in RNA-directed DNA methylation (RdDM) in plants. Here, we provide evidence, based on laser UV-assisted zero-length... more
RNA polymerase V (Pol V) long noncoding RNAs (lncRNAs) have been proposed to guide ARGONAUTE4 (AGO4) to chromatin in RNA-directed DNA methylation (RdDM) in plants. Here, we provide evidence, based on laser UV-assisted zero-length cross-linking, for functionally relevant AGO4-DNA interaction at RdDM targets. We further demonstrate that Pol V lncRNAs or the act of their transcription are required to lock Pol V holoenzyme into a stable DNA-bound state that allows AGO4 recruitment via redundant glycine-tryptophan/tryptophan-glycine AGO hook motifs present on both Pol V and its associated factor, SPT5L. We propose a model in which AGO4-DNA interaction could be responsible for the unique specificities of RdDM.
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Genomic DNA is prone to a large number of insults by a myriad of endogenous and exogenous agents. The base excision repair (BER) is the major mechanism used by cells for the removal of various DNA lesions spontaneously or environmentally... more
Genomic DNA is prone to a large number of insults by a myriad of endogenous and exogenous agents. The base excision repair (BER) is the major mechanism used by cells for the removal of various DNA lesions spontaneously or environmentally induced and the maintenance of genome integrity. The presence of persistent DNA damage is not compatible with life, since abrogation of BER leads to early embryonic lethality in mice. There are several lines of evidences showing existence of a link between deficient BER, cancer proneness and ageing, thus illustrating the importance of this DNA repair pathway in human health. Although the enzymology of BER mechanisms has been largely elucidated using chemically defined DNA damage substrates and purified proteins, the complex interplay of BER with another vital process like transcription or when DNA is in its natural state (i.e. wrapped in nucleosome and assembled in chromatin fiber is largely unexplored. Cells use chromatin remodeling factors to over...
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FACT, in addition to its role in transcription, is likely implicated in both transcription-coupled nucleotide excision repair and DNA double strand break repair. Here, we present evidence that FACT could be directly involved in Base... more
FACT, in addition to its role in transcription, is likely implicated in both transcription-coupled nucleotide excision repair and DNA double strand break repair. Here, we present evidence that FACT could be directly involved in Base Excision Repair and elucidate the chromatin remodeling mechanisms of FACT during BER. We found that, upon oxidative stress, FACT is released from transcription related protein complexes to get associated with repair proteins and chromatin remodelers from the SWI/SNF family. We also showed the rapid recruitment of FACT to the site of damage, coincident with the glycosylase OGG1, upon the local generation of oxidized DNA. Interestingly, FACT facilitates uracil-DNA glycosylase in the removal of uracil from nucleosomal DNA thanks to an enhancement in the remodeling activity of RSC. This discloses a novel property of FACT wherein it has a co-remodeling activity and strongly enhances the remodeling capacity of the chromatin remodelers. Altogether, our data sug...
Research Interests: Genetics, Oxidative Stress, DNA damage, DNA repair, Transcription Factors, and 15 moreSaccharomyces cerevisiae, Humans, Chromatin Remodeling and Epigenetics, Animals, Chromatin remodeling, Nucleosomes, Chromatin, Histones, HeLa cells, Xenopus laevis, PLoS Genetics, Fact, DNA binding proteins, High mobility group proteins, and Uracil
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Research Interests:
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Thyroid hormones control various aspects of gut development and homeostasis. The best-known example is in gastrointestinal tract remodeling during amphibian metamorphosis. It is well documented that these hormones act via the TR nuclear... more
Thyroid hormones control various aspects of gut development and homeostasis. The best-known example is in gastrointestinal tract remodeling during amphibian metamorphosis. It is well documented that these hormones act via the TR nuclear receptors, which are hormone-modulated transcription factors. Several studies have shown that thyroid hormones regulate the expression of several genes in the Notch signaling pathway, indicating a possible means by which they participate in the control of gut physiology. However, the mechanisms and biological significance of this control have remained unexplored. Using multiple in vivo and in vitro approaches, we show that thyroid hormones positively regulate Notch activity through the TRα1 receptor. From a molecular point of view, TRα1 indirectly controls Notch1, Dll1, Dll4 and Hes1 expression but acts as a direct transcriptional regulator of the Jag1 gene by binding to a responsive element in the Jag1 promoter. Our findings show that the TRα1 nucle...
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Clustered DNA damage is a unique characteristic of radiation-induced DNA damage and the formation of these sites poses a serious challenge to the... more
Clustered DNA damage is a unique characteristic of radiation-induced DNA damage and the formation of these sites poses a serious challenge to the cell's repair machinery. Within a cell DNA is compacted, with nucleosomes being the first order of higher level structure. However, few data are reported on the efficiency of clustered-lesion processing within nucleosomal DNA templates. Here, we show retardation of cleavage of a single AP site by purified APE1 when contained in nucleosomal DNA, compared to cleavage of an AP site in non-nucleosomal DNA. This retardation seen in nucleosomal DNA was alleviated by incubation with CHO-K1 nuclear extract. When clustered DNA damage sites containing bistranded AP sites were present in nucleosomal DNA, efficient cleavage of the AP sites was observed after treatment with nuclear extract. The resultant DSB formation led to DNA dissociating from the histone core and nucleosomal dispersion. Clustered damaged sites containing bistranded AP site/8-oxoG residues showed no retardation of cleavage of the AP site but retardation of 8-oxoG excision, compared to isolated lesions, thus DSB formation was not seen. An increased understanding of processing of clustered DNA damage in a nucleosomal environment may lead to new strategies to enhance the cytotoxic effects of radiotherapeutics.
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A laser photoionization setup is presented for the study of highly excited Rydberg and autoionization atomic states. The setup includes three dye lasers pumped by an N2 laser and is designed for spectroscopic and analytical purposes. The... more
A laser photoionization setup is presented for the study of highly excited Rydberg and autoionization atomic states. The setup includes three dye lasers pumped by an N2 laser and is designed for spectroscopic and analytical purposes. The spectrometer has a spectral resolution of greater than or equal to 0.8/cm, making it possible to accurately determine the energies and widths of highly excited atomic states. The present photoionization setup can be used as an ultrasensitive analyzer of elements in various solid or liquid samples without any preliminary chemical treatment.
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ABSTRACT The first successful experiments on selective interaction with a specific type of base in DNA and RNA are reported. It is shown that when dilute aqueous solutions of bases are irradiated by high-power ultrashort ultraviolet laser... more
ABSTRACT The first successful experiments on selective interaction with a specific type of base in DNA and RNA are reported. It is shown that when dilute aqueous solutions of bases are irradiated by high-power ultrashort ultraviolet laser pulses, two-stage excitation of high electronic levels takes place, resulting in the formation of irreversible photoproducts. The quantum yield of photoproducts depends on many parameters and may be made essentially different for bases of different types. By varying the experimental conditions, selective interaction with a specific type of base was achieved both by irradiating mixtures of bases and by irradiating more complex components of nucleic acids.
Generation, in a KDP crystal, of the sum frequencies of YAG:Nd3+ laser harmonics and the Stokes components of stimulated Raman scattering produced picosecond pulses of 218–316 nm wavelengths and 0.1–3 mJ energy.
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Nanosecond laser-induced oxidative lesions at guanines within DNA were investigated using sequencing gel electrophoresis analysis. Strong sequence-specific lesions on guanines were revealed by either Fpg or piperidine treatments and... more
Nanosecond laser-induced oxidative lesions at guanines within DNA were investigated using sequencing gel electrophoresis analysis. Strong sequence-specific lesions on guanines were revealed by either Fpg or piperidine treatments and assigned to 8-oxoG and oxazolone, respectively. It was shown that both the biphotonic ionization process and the chemical reactivity of the radical cation (G+) are dependent on the DNA sequence. The former were explained by the occurrence of energy and charge migration phenomena, while the latter in terms of local DNA hydration peculiarities.
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The rare base 4-thiouridine (s4U), present in various transfer RNA (tRNA) molecules from Escherichia coli, occupies usually the strategically important 8th position between the double helices of the acceptor and the dihydrouridine stems... more
The rare base 4-thiouridine (s4U), present in various transfer RNA (tRNA) molecules from Escherichia coli, occupies usually the strategically important 8th position between the double helices of the acceptor and the dihydrouridine stems of the cloverleaf. This unusual base is largely used as an intrinsic build-in probe for RNA conformational and RNA(DNA)-protein interaction studies through triplet excited state photochemistry related to covalent adducts formation. Here, by applying laser transient absorption saturation spectroscopy, we measured the intersystem crossing yield jST and the excited triplet state absorption eT-T of s4U within tRNA. While the incorporation of s4U in tRNA induced appreciable changes in the latter, no important variation of the intersystem crossing yield was observed, which is in contrasts with the published data.
ABSTRACT A set-up including three dye lasers pumped by a N2-laser for selective multistep ionization of atomic beam is described. The experiment is designed not only for spectroscopic investigation but for analytical use too. The... more
ABSTRACT A set-up including three dye lasers pumped by a N2-laser for selective multistep ionization of atomic beam is described. The experiment is designed not only for spectroscopic investigation but for analytical use too. The application of the described set-up is briefly discussed. Some results on experimentally investigated highly-excited Rydberg states of Thulium are reported. Model potential calculations allow correct assignment of the spectra to be carried out. Non-hydrogen behaviour of the Rydberg states of rare-earth atoms is discussed.
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In the first part of the chapter, emphasis is placed on the description of the main reactions of radical cations of the four predominant DNA purine and pyrimidine bases and minor 5-methylcytosine in aerated aqueous solutions. Information... more
In the first part of the chapter, emphasis is placed on the description of the main reactions of radical cations of the four predominant DNA purine and pyrimidine bases and minor 5-methylcytosine in aerated aqueous solutions. Information is also provided on the final one-electron oxidation products of 8-oxo-7,8-dihydroguanine, an ubiquitous decomposition product of DNA with most chemical and physical oxidizing
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Mammalian centromeric histone H3 variant, CENP-A, is involved in maintaining the functional integrity and epigenetic inheritance of the centromere. CENP-A causes transcriptional repression of centromeric chromatin through an unknown... more
Mammalian centromeric histone H3 variant, CENP-A, is involved in maintaining the functional integrity and epigenetic inheritance of the centromere. CENP-A causes transcriptional repression of centromeric chromatin through an unknown mechanism. Here, we report that reconstituted CENP-A nucleosomes are amenable to ATP-dependent SWI/SNF-mediated remodelling but are less permissive to acetylation and acetylation-dependent in vitro chromatin transcription. Remarkably, the transcriptional repression of the CENP-A chromatinized template could be relieved by the ectopic addition of histone chaperone, nucleophosmin.
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In this work we have studied the mechanisms of regulation of expression of androgen receptor (AR) target genes. We have used an immortalized non-tumorigenic prostate cell line RWPE-1-AR(tag) constitutively expressing an exogenous AR as a... more
In this work we have studied the mechanisms of regulation of expression of androgen receptor (AR) target genes. We have used an immortalized non-tumorigenic prostate cell line RWPE-1-AR(tag) constitutively expressing an exogenous AR as a model. We observed that all studied AR target genes exhibited a specific expression during the G1 phase of the cell cycle despite the constitutive expression of AR. Importantly, we found that the expression of NCoR, an AR co-repressor, was downregulated during the G1 phase and expressed as mRNA and protein specifically during the S phase. The role of NCoR in repressing androgen-induced expression of AR target genes in S phase was further demonstrated by altering expression of NCoR during the cell cycle through knockdown or induced overexpression. Using two alternative techniques we show that AR binds directly to target DNA in the chromatin only during the G1 phase. These data support the hypothesis that NCoR might control a cell cycle dependent regu...
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Within chromatin, the core histone tail domains play critical roles in regulating the structure and accessibility of nucleosomal DNA within the chromatin fiber. Thus, many nuclear processes are facilitated by concomitant posttranslational... more
Within chromatin, the core histone tail domains play critical roles in regulating the structure and accessibility of nucleosomal DNA within the chromatin fiber. Thus, many nuclear processes are facilitated by concomitant posttranslational modification of these domains. However, elucidation of the mechanisms by which the tails mediate such processes awaits definition of tail interactions within chromatin. In this study we have investigated the primary DNA target of the majority of the tails in mononucleosomes. The results clearly show that the tails bind preferentially to "linker" DNA, outside of the DNA encompassed by the nucleosome core. These results have important implications for models of tail function within the chromatin fiber and for in vitro structural and functional studies using nucleosome core particles.
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ABSTRACT The photoinduced effects on nucleic acids and some of their components by high-power laser are compared to those observed under the influence of high energy irradiation (electrons and γ-rays). Similarities of the action of both... more
ABSTRACT The photoinduced effects on nucleic acids and some of their components by high-power laser are compared to those observed under the influence of high energy irradiation (electrons and γ-rays). Similarities of the action of both kinds of radiation and the resulting transients are discussed on the basis of some model compounds.
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Two cell repair systems--photoreactivation and repair of single-strand DNA breaks have been studied using unicellular green algae as a test-system. Effects of the genotype and the intensity of pico/second UV-laser irradiation on the... more
Two cell repair systems--photoreactivation and repair of single-strand DNA breaks have been studied using unicellular green algae as a test-system. Effects of the genotype and the intensity of pico/second UV-laser irradiation on the degree of the photoreactivation have been investigated. It has been shown that the lower intensity (I = 8.10(6) W/cm2) effects less the inactivation of living cells comparing with I = 30.10(6) W/cm2, regardless of the genotype. The clearly expressed higher potentials of strains LARG-1 and 260 to produce and repair alterations of the cyclobutane-pyrimidine dimers type have been established. An analysis of DNA degradation during gamma rays irradiation and after incubation has been carried out for investigation the relationship between strains radioresistance and repair of single-strand break. It has been shown that high efficiency of the repair system is characteristic of the resistant strain obtained from chronically irradiated population.
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... Fedoseev, V.; Ivanov, LN. Publication: Laser Spectroscopy VI; Proceedings of the Sixth International Conference, held June 27 - July 1, 1983, in Interlaken, Switzerland. Edited by HP Weber and W. Lüthy. Volume 40 of the Springer... more
... Fedoseev, V.; Ivanov, LN. Publication: Laser Spectroscopy VI; Proceedings of the Sixth International Conference, held June 27 - July 1, 1983, in Interlaken, Switzerland. Edited by HP Weber and W. Lüthy. Volume 40 of the Springer Series in Optical Sciences, edited by Arthur L ...
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ABSTRACT A picosecond UV laser radiation was used to cross-link proteins to DNA in nuclei, whole cells and different chromating preparations. All histones as well as high-mobility group 1 proten were identified immunochemically in the... more
ABSTRACT A picosecond UV laser radiation was used to cross-link proteins to DNA in nuclei, whole cells and different chromating preparations. All histones as well as high-mobility group 1 proten were identified immunochemically in the covalently linked protein-DNA complexes. Irradiation of the nuclieohistone resulted in cross-linking 20% of bound histones to DNA as a result of two-quantum photoreaction with a maximum quantum yield 3.10 -4 for double stranded DNA. When nuclei, total bromatin Hi-depleted chromatin and core particles were irradiated and then trypsinized or treated with clostripain to cleave respectively the N-, C- and N- terminal histone tails, no histones have been found covalently linked to DNA. However whilst the yield of cross-links was similar in total and H1-depleted chromatin in core particples the efficiencey was 3-4 times lower for H2A, H2B and H4 10-12 times lower for H3. This finding we consider as a direct evidence for interaction of non structured N- tails of core histones with linker DNA. Cross-linking in core particles depends on the ionic strength. All histones were identified in the complex formed up to 0.4 N NaCl, no cross-linking was observed when irradiation was carried out at salt concentration higher than 0.4 M. The cross-linking ability was preserved both upon physiological acetylation of histones knows to be restriced to the N-terminal tails and with chemically acetylated chromatin. This finding is direct evidence that postsynthetic histone acetylation does not release the N-terminal tails from interaction with DNA.
ABSTRACT
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Research Interests: Cellular Biology, Gene expression, Biological Sciences, Cell Differentiation, Molecular and cellular biology, and 13 moreHumans, Mice, Animals, Retinoblastoma, Histones, Rats, Adult, Base Sequence, Gene Expression Regulation, retinoblastoma protein (RB1)., Molecular Sequence Data, High mobility group proteins, and Medical and Health Sciences
ABSTRACT A comparative experimental study of two versions of an injection-seeding laser system containing a slave laser (SL) with a ring-linear dual-channel competitive cavity and with a simple ring one is presented. The spectral purity... more
ABSTRACT A comparative experimental study of two versions of an injection-seeding laser system containing a slave laser (SL) with a ring-linear dual-channel competitive cavity and with a simple ring one is presented. The spectral purity of the SL output radiation and efficiency of the SL reverse wave suppression in dependence on the seeding conditions are investigated. It is shown that in the case of a dual-cavity competitive SL with a ring main channel both the spectral background and reverse radiation are fully suppressed. This is realized in a compact scheme without optical isolators and at any ratios between the SL energy and the seeding source one.
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ABSTRACT A laser cavity containing highly selective reflector based on a multipass grazing-incidence diffraction grating is described. The operation of the multipass selective reflector (MPSR) is demonstrated and studied experimentally in... more
ABSTRACT A laser cavity containing highly selective reflector based on a multipass grazing-incidence diffraction grating is described. The operation of the multipass selective reflector (MPSR) is demonstrated and studied experimentally in the case of pulsed dye laser pumped by frequency-doubled Nd:YAG laser. Numerical calculations concerning gain- switched Ti:Sapphire laser with MPSR are also done. An increasing of the spectrally pure output energy more than 5 times is achieved. Decrease of the threshold energy and broadening of the tuning range of several times are also observed depending on the pumping conditions. The calculations show that, in the case of gain-switched Ti:Sapphire laser pumped by Nd:YAG laser, the temporal characteristics of the radiation, the build-up time and the pulse duration are considerably shortened by using MPSR. The improved energy, spectral, and temporal parameters of tunable lasers with MPSR make them suitable for different applications (spectroscopy, biology, DIAL, etc.) as a separate sources or as a part of injection-seeding tunable laser systems.
ABSTRACT An injection seeding of narrowband and broadband radiation into a highly selective tunable laser is described. An increase of the laser output energy by a factor of 1,5 is achieved. Laser oscillations are induced by injection... more
ABSTRACT An injection seeding of narrowband and broadband radiation into a highly selective tunable laser is described. An increase of the laser output energy by a factor of 1,5 is achieved. Laser oscillations are induced by injection seeding when the active medium is pumped below the laser threshold. Optical signal as small as 0.2 nJ is detected and amplified by a factor of 150.
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Light-induced absorption in pure B12TiO20 crystal and crystals doped with Nd, Mo, W, and V was investigated using the pump-probe method. Q-switched frequency-doubled (532 nm) YAG:Nd laser with pulse duration approximately 20 ns was... more
Light-induced absorption in pure B12TiO20 crystal and crystals doped with Nd, Mo, W, and V was investigated using the pump-probe method. Q-switched frequency-doubled (532 nm) YAG:Nd laser with pulse duration approximately 20 ns was applied for sample irradiation. Investigation of pump-induced transmission was carried out by means of a tunable dye laser radiation at (lambda) 1 equals 568 nm and (lambda) 2 equals 698 nm. The dependence of the light-induced absorption coefficient on the pump beam intensity and probe wavelengths was determined. The two-center model was used to explain the observed results.