Loss of prohibitin 2 in Schwann cells dysregulates key transcription factors controlling developmental myelination.

Schwann cells are critical for the proper development and function of the peripheral nervous system, where they form a mutually beneficial relationship with axons. Past studies have highlighted that a pair of proteins called the prohibitins play major roles in Schwann cell biology. Prohibitins are ubiquitously expressed and versatile proteins. We have previously shown that while prohibitins play a crucial role in Schwann cell mitochondria for long-term myelin maintenance and axon health, they may also be present at the Schwann cell-axon interface during development. Here, we expand on this work, showing that drug-mediated modulation of prohibitins in vitro disrupts myelination and confirming that Schwann cell-specific ablation of prohibitin 2 (Phb2) in vivo results in early and severe defects in peripheral nerve development. Using a proteomic approach in vitro, we identify a pool of candidate PHB2 interactors that change their interaction with PHB2 depending on the presence of axonal signals. Furthermore, we show in vivo that loss of Phb2 in mouse Schwann cells causes ineffective proliferation and dysregulation of transcription factors EGR2 (KROX20), POU3F1 (OCT6) and POU3F2 (BRN2) that are necessary for proper Schwann cell maturation. Schwann cell-specific deletion of Jun, a transcription factor associated with negative regulation of myelination, confers partial rescue of the development defect seen in mice lacking Schwann cell Phb2. This work develops our understanding of Schwann cell biology, revealing that Phb2 may directly or indirectly modulate the timely expression of transcription factors necessary for proper peripheral nervous system development, and proposing candidates that may play a role in PHB2-mediated integration of axon signals in the Schwann cell.

Transcription Factors in Brain Regeneration: A Potential Novel Therapeutic Target.

Transcription factors play a crucial role in providing identity to each cell population. To maintain cell identity, it is essential to balance the expression of activator and inhibitor transcription factors. Cell plasticity and reprogramming offer great potential for future therapeutic applications, as they can regenerate damaged tissue. Specific niche factors can modify gene expression and differentiate or transdifferentiate the target cell to the required fate. Ongoing research is being carried out on the possibilities of transcription factors in regenerating neurons, with neural stem cells (NSCs) being considered the preferred cells for generating new neurons due to their epigenomic and transcriptome memory. NEUROD1/ASCL1, BRN2, MYTL1, and other transcription factors can induce direct reprogramming of somatic cells, such as fibroblasts, into neurons. However, the molecular biology of transcription factors in reprogramming and differentiation still needs to be fully understood.

Peptide L13S Derived from the BRN2 POU Domain Reduces Metastasis In Vivo and Inhibits Cell Migration and Invasion In Vitro.

BACKGROUND/AIM: The Brain-Specific Homeobox/POU Domain Protein 2 (BRN2) transcription factor supports melanoma progression by regulating the expression of several genes involved in cell migration and invasion. We hypothesized that a peptide designed based on the POU domain of BRN2 could block the BRN2 transcription activity and, consequently, reduce metastasis. MATERIALS AND METHODS: Cell viability was accessed by Trypan Blue exclusion dye assay and xCelligence platform. Wound-healing scratch assay and transwell invasion with matrigel membrane assay were performed to analyze cell migration and invasion. The internalization mechanism of the L13S peptide was investigated using confocal microscopy and wound-healing scratch assay. The impact of L13S on cell protein expression was analyzed through western blotting. In vivo assays were conducted to evaluate the protective effect and toxicity of L13S in a metastatic model using murine melanoma cells. RESULTS: Here, we show that the peptide named L13S can inhibit the migration and invasion of murine melanoma cells (B16F10-Nex2) as well as the migration of human melanoma cells (SK-MEL-25 and A375) by regulating the expression of proteins involved in motility. Mechanistically, we found that L13S is internalized by murine melanoma cells via macropinocytosis and binds actin filaments and nuclei. More importantly, in vivo studies indicated that the peptide was able to significantly inhibit lung metastasis in syngeneic models without off-target effects and with virtually no cytotoxicity toward normal organs. CONCLUSION: L13S peptide is a strong candidate for further development as an anticancer agent for the treatment of melanoma metastasis.

Monoallelic intragenic POU3F2 variants lead to neurodevelopmental delay and hyperphagic obesity, confirming the gene's candidacy in 6q16.1 deletions.

While common obesity accounts for an increasing global health burden, its monogenic forms have taught us underlying mechanisms via more than 20 single-gene disorders. Among these, the most common mechanism is central nervous system dysregulation of food intake and satiety, often accompanied by neurodevelopmental delay (NDD) and autism spectrum disorder. In a family with syndromic obesity, we identified a monoallelic truncating variant in POU3F2 (alias BRN2) encoding a neural transcription factor, which has previously been suggested as a driver of obesity and NDD in individuals with the 6q16.1 deletion. In an international collaboration, we identified ultra-rare truncating and missense variants in another ten individuals sharing autism spectrum disorder, NDD, and adolescent-onset obesity. Affected individuals presented with low-to-normal birth weight and infantile feeding difficulties but developed insulin resistance and hyperphagia during childhood. Except for a variant leading to early truncation of the protein, identified variants showed adequate nuclear translocation but overall disturbed DNA-binding ability and promotor activation. In a cohort with common non-syndromic obesity, we independently observed a negative correlation of POU3F2 gene expression with BMI, suggesting a role beyond monogenic obesity. In summary, we propose deleterious intragenic variants of POU3F2 to cause transcriptional dysregulation associated with hyperphagic obesity of adolescent onset with variable NDD.

The Transcription Factor Pou3f1 Sheds Light on the Development and Molecular Diversity of Glutamatergic Cerebellar Nuclear Neurons in the Mouse.

The cerebellar nuclear (CN) neurons are a molecularly heterogeneous population whose specification into the different cerebellar nuclei is defined by the expression of varying sets of transcription factors. Here, we present a novel molecular marker, Pou3f1, that delineates specific sets of glutamatergic CN neurons. The glutamatergic identity of Pou3f1+ cells was confirmed by: (1) the co-expression of vGluT2, a cell marker of glutamatergic neurons; (2) the lack of co-expression between Pou3f1 and GAD67, a marker of GABAergic neurons; (3) the co-expression of Atoh1, the master regulator required for the production of all cerebellar glutamatergic lineages; and (4) the absence of Pou3f1-expressing cells in the Atoh1-null cerebellum. Furthermore, the lack of Pax6 and Tbr1 expression in Pou3f1+ cells reveals that Pou3f1-expressing CN neurons specifically settle in the interposed and dentate nuclei. In addition, the Pou3f1-labeled glutamatergic CN neurons can be further classified by the expression of Brn2 and Irx3. The results of the present study align with previous findings highlighting that the survival of the interposed and dentate CN neurons is largely independent of Pax6. More importantly, the present study extends the field's collective knowledge of the molecular diversity of cerebellar nuclei.

[Transcription Factors of Direct Neuronal Reprogramming in Ontogenesis and ex vivo].

Direct reprogramming technology allows several specific types of cells, including specialized neurons, to be obtained from readily available autologous somatic cells. It presents unique opportunities for the development of personalized medicine, from in vitro models of hereditary and degenerative neurological diseases to novel neuroregenerative technologies. Over the past decade, a plethora of protocols for primary reprogramming has been published, yet reproducible generation of homogeneous populations of neuronally reprogrammed cells still remains a challenge. All existing protocols, however, use transcription factors that are involved in embryonic neurogenesis. This is presumably be the key issue for obtaining highly efficient and reproducible protocols for ex vivo neurogenesis. Analysis of the functional features of transcription factors in embryonic and adult neurogenesis may not only lead to the improvement of reprogramming protocols, but also, via cell marker analysis, can exactly determine the stage of neurogenesis that a particular protocol will reach. The purpose of this review is to characterize the general factors that play key roles in neurogenesis for the embryonic and adult periods, as well as in cellular reprogramming, and to assess correspondence of cell forms obtained as a result of cellular reprogramming to the ontogenetic series of the nervous system, from pluripotent stem cells to specialized neurons.

Isolation and Neuronal Reprogramming of Mouse Embryonic Fibroblasts.

Forced expression of specific neuronal transcription factors in mouse embryonic fibroblasts (MEFs) can lead to their direct conversion into functional neurons. Direct neuronal reprogramming has become a powerful tool to characterize individual factors and molecular mechanisms involved in forced and normal neurogenesis and to generate neuronal cell types for in vitro studies. Here we provide a detailed protocol for the isolation of MEFs devoid of neural tissue and their direct reprogramming into functional neurons by overexpression of neuronal reprogramming factors (Ascl1, Brn2, and Myt1l) using lentiviral vectors. This method enables quick and efficient generation of mouse neurons in vitro for versatile functional and mechanistic characterization.

Hierarchical reactivation of transcription during mitosis-to-G1 transition by Brn2 and Ascl1 in neural stem cells.

During mitosis, chromatin condensation is accompanied by a global arrest of transcription. Recent studies suggest transcriptional reactivation upon mitotic exit occurs in temporally coordinated waves, but the underlying regulatory principles have yet to be elucidated. In particular, the contribution of sequence-specific transcription factors (TFs) remains poorly understood. Here we report that Brn2, an important regulator of neural stem cell identity, associates with condensed chromatin throughout cell division, as assessed by live-cell imaging of proliferating neural stem cells. In contrast, the neuronal fate determinant Ascl1 dissociates from mitotic chromosomes. ChIP-seq analysis reveals that Brn2 mitotic chromosome binding does not result in sequence-specific interactions prior to mitotic exit, relying mostly on electrostatic forces. Nevertheless, surveying active transcription using single-molecule RNA-FISH against immature transcripts reveals differential reactivation kinetics for key targets of Brn2 and Ascl1, with transcription onset detected in early (anaphase) versus late (early G1) phases, respectively. Moreover, by using a mitotic-specific dominant-negative approach, we show that competing with Brn2 binding during mitotic exit reduces the transcription of its target gene Nestin Our study shows an important role for differential binding of TFs to mitotic chromosomes, governed by their electrostatic properties, in defining the temporal order of transcriptional reactivation during mitosis-to-G1 transition.

Integrin α3ß1 Promotes Invasive and Metastatic Properties of Breast Cancer Cells through Induction of the Brn-2 Transcription Factor.

In the current study, we demonstrate that integrin α3ß1 promotes invasive and metastatic traits of triple-negative breast cancer (TNBC) cells through induction of the transcription factor, Brain-2 (Brn-2). We show that RNAi-mediated suppression of α3ß1 in MDA-MB-231 cells caused reduced expression of Brn-2 mRNA and protein and reduced activity of the BRN2 gene promoter. In addition, RNAi-targeting of Brn-2 in MDA-MB-231 cells decreased invasion in vitro and lung colonization in vivo, and exogenous Brn-2 expression partially restored invasion to cells in which α3ß1 was suppressed. α3ß1 promoted phosphorylation of Akt in MDA-MB-231 cells, and treatment of these cells with a pharmacological Akt inhibitor (MK-2206) reduced both Brn-2 expression and cell invasion, indicating that α3ß1-Akt signaling contributes to Brn-2 induction. Analysis of RNAseq data from patients with invasive breast carcinoma revealed that high BRN2 expression correlates with poor survival. Moreover, high BRN2 expression positively correlates with high ITGA3 expression in basal-like breast cancer, which is consistent with our experimental findings that α3ß1 induces Brn-2 in TNBC cells. Together, our study demonstrates a pro-invasive/pro-metastatic role for Brn-2 in breast cancer cells and identifies a role for integrin α3ß1 in regulating Brn-2 expression, thereby revealing a novel mechanism of integrin-dependent breast cancer cell invasion.

Loss of CHD1 Promotes Heterogeneous Mechanisms of Resistance to AR-Targeted Therapy via Chromatin Dysregulation.

Metastatic prostate cancer is characterized by recurrent genomic copy number alterations that are presumed to contribute to resistance to hormone therapy. We identified CHD1 loss as a cause of antiandrogen resistance in an in vivo small hairpin RNA (shRNA) screen of 730 genes deleted in prostate cancer. ATAC-seq and RNA-seq analyses showed that CHD1 loss resulted in global changes in open and closed chromatin with associated transcriptomic changes. Integrative analysis of this data, together with CRISPR-based functional screening, identified four transcription factors (NR3C1, POU3F2, NR2F1, and TBX2) that contribute to antiandrogen resistance, with associated activation of non-luminal lineage programs. Thus, CHD1 loss results in chromatin dysregulation, thereby establishing a state of transcriptional plasticity that enables the emergence of antiandrogen resistance through heterogeneous mechanisms.

BRN2 suppresses apoptosis, reprograms DNA damage repair, and is associated with a high somatic mutation burden in melanoma.

Whether cell types exposed to a high level of environmental insults possess cell type-specific prosurvival mechanisms or enhanced DNA damage repair capacity is not well understood. BRN2 is a tissue-restricted POU domain transcription factor implicated in neural development and several cancers. In melanoma, BRN2 plays a key role in promoting invasion and regulating proliferation. Here we found, surprisingly, that rather than interacting with transcription cofactors, BRN2 is instead associated with DNA damage response proteins and directly binds PARP1 and Ku70/Ku80. Rapid PARP1-dependent BRN2 association with sites of DNA damage facilitates recruitment of Ku80 and reprograms DNA damage repair by promoting Ku-dependent nonhomologous end-joining (NHEJ) at the expense of homologous recombination. BRN2 also suppresses an apoptosis-associated gene expression program to protect against UVB-, chemotherapy- and vemurafenib-induced apoptosis. Remarkably, BRN2 expression also correlates with a high single-nucleotide variation prevalence in human melanomas. By promoting error-prone DNA damage repair via NHEJ and suppressing apoptosis of damaged cells, our results suggest that BRN2 contributes to the generation of melanomas with a high mutation burden. Our findings highlight a novel role for a key transcription factor in reprogramming DNA damage repair and suggest that BRN2 may impact the response to DNA-damaging agents in BRN2-expressing cancers.

A PAX3/BRN2 rheostat controls the dynamics of BRAF mediated MITF regulation in MITFhigh /AXLlow melanoma.

The BRAF kinase and the MAPK pathway are targets of current melanoma therapies. However, MAPK pathway inhibition results in dynamic changes of downstream targets that can counteract inhibitor-action not only in during treatment, but also in acquired resistant tumours. One such dynamic change involves the expression of the transcription factor MITF, a crucial regulator of cell survival and proliferation in untreated as well as drug-addicted acquired resistant melanoma. Tight control over MITF expression levels is required for optimal melanoma growth, and while it is well established that the MAPK pathway regulates MITF expression, the actual mechanism is insufficiently understood. We reveal here, how BRAF through action on the transcription factors BRN2 and PAX3 executes control over the regulation of MITF expression in a manner that allows for considerable plasticity. This plasticity provides robustness to the BRAF mediated MITF regulation and explains the dynamics in MITF expression that are observed in patients in response to MAPK inhibitor therapy.

Peptide R18H from BRN2 Transcription Factor POU Domain Displays Antitumor Activity In Vitro and In Vivo and Induces Apoptosis in B16F10-Nex2 Cells.

BACKGROUND: BRN2 transcription factor is associated with the development of malignant melanoma. The cytotoxic activities and cell death mechanism against B16F10-Nex2 cells were determined with synthetic peptide R18H derived from the POU domain of the BRN2 transcription factor. OBJECTIVE: To determine the cell death mechanisms and in vivo activity of peptide R18H derived from the POU domain of the BRN2 transcription factor against B16F10-Nex2 cells. METHODS: Cell viability was determined by the MTT method. C57Bl/6 mice were challenged with B16F10-Nex2 cells and treated with R18H. To identify the type of cell death, we used TUNEL assay, Annexin V and PI, Hoechst, DHE, and determination of caspase activation and cytochrome c release. Transmission electron microscopy was performed to verify morphological alterations after peptide treatment. RESULTS: Peptide R18H displayed antitumor activity in the first hours of treatment and the EC50% was calculated for 2 and 24h, being 0.76 ± 0.045 mM and 0.559 ± 0.053 mM, respectively. After 24h apoptosis was evident, based on DNA degradation, chromatin condensation, increase of superoxide anion production, phosphatidylserine translocation, activation of caspases 3 and 8, and release of extracellular cytochrome c in B16F10-Nex2 cells. The peptide cytotoxic activity was not affected by necroptosis inhibitors and treated cells did not release LDH in the extracellular medium. Moreover, in vivo antitumor activity was observed following treatment with peptide R18H. CONCLUSION: Peptide R18H from BRN2 transcription factor induced apoptosis in B16F10-Nex2 and displayed antitumor activity in vivo.

Bi-allelic Loss of CDKN2A Initiates Melanoma Invasion via BRN2 Activation.

Loss of the CDKN2A tumor suppressor is associated with melanoma metastasis, but the mechanisms connecting the phenomena are unknown. Using CRISPR-Cas9 to engineer a cellular model of melanoma initiation from primary human melanocytes, we discovered that a lineage-restricted transcription factor, BRN2, is downstream of CDKN2A and directly regulated by E2F1. In a cohort of melanocytic tumors that capture distinct progression stages, we observed that CDKN2A loss coincides with both the onset of invasive behavior and increased BRN2 expression. Loss of the CDKN2A protein product p16INK4A permitted metastatic dissemination of human melanoma lines in mice, a phenotype rescued by inhibition of BRN2. These results demonstrate a mechanism by which CDKN2A suppresses the initiation of melanoma invasion through inhibition of BRN2.

Brn2 Alone Is Sufficient to Convert Astrocytes into Neural Progenitors and Neurons.

Generating neurons or neural progenitor cells (NPCs) from astrocytes is a potential strategy for neurological repair by reprogramming. Previous study has showed that Brn2, by cooperating with other factors, participates in neurogenesis and neuronal reprogramming. However, it is still unclear whether the Brn2 alone can convert astrocytes into neurons or NPCs. Here, we explored the effect of Brn2 on reprogramming of astrocytes, and found that a single transcription factor Brn2 can convert mouse astrocytes into functional neurons. Furthermore, the Brn2-infected astrocytes can be induced into NPCs after changing culture condition. In addition, our study found that the reprogramming of astrocytes and the fate of transdifferentiated cells are closely associated with cell microenvironmental factors, such as the brain regions where the astrocytes come from, the proliferation ability of astrocytes, and culture condition of infected astrocytes. To sum up, for the first time, our results demonstrated that Brn2 alone is sufficient to convert astrocytes into neural progenitors and neurons, and the conversion is associated with cell microenvironments. This new conversion method will be a potential therapeutic approach to restore the injured diseased brain in regenerative medicine.

POU3F2 participates in cognitive function and adult hippocampal neurogenesis via mammalian-characteristic amino acid repeats.

POU3F2/BRN-2 is a transcription factor that is mainly expressed in the central nervous system and plays an important role in brain development. The transactivation domain of POU3F2 includes multiple mammalian-characteristic tandem amino acid repeats (homopolymeric amino acid repeats). We previously generated knock-in mice (Pou3f2Δ/Δ mice) in which all three homopolymeric amino acid repeats were deleted from the Pou3f2 transactivation domain and identified phenotypic impairments in maternal behavior and pup recognition. Yet, the exact biological implications of homopolymeric repeats are not completely understood. In this study, we investigated cognitive function and hippocampal neurogenesis in Pou3f2Δ/Δ mice. Pou3f2Δ/Δ mice exhibited cognitive impairment in object recognition and object location tests. Immunohistochemistry for doublecortin, a marker of immature neurons, showed a lower number of newborn neurons in the dentate gyrus of adult Pou3f2Δ/Δ mice compared with wild-type mice. Consistent with this observation, adult Pou3f2Δ/Δ mice had lower numbers of 5-bromo-2'-deoxyuridine (BrdU) and NeuN double-positive cells at 4 weeks after BrdU injection compared with control mice, indicating the decreased generation of mature granule cells in Pou3f2Δ/Δ mice. Taken together, these results suggest that POU3F2 is involved in cognitive function as well as adult hippocampal neurogenesis, and that homopolymeric amino acid repeats in this gene play a functional role.

Effects of Brn2 overexpression on eccrine sweat gland development in the mouse paw.

Eccrine sweat glands regulate body temperature by secreting water and electrolytes. In humans, eccrine sweat glands are ubiquitous in the skin, except in the lips and external genitalia. In mice, eccrine sweat glands are present only in the paw pad. Brn2 is a protein belonging to a large family of transcription factors. A few studies have examined Brn2 in melanoma cells and epidermal keratinocytes. This study investigated changes in the skin in the K5-Brn2 transgenic mouse, which overexpresses Brn2 and contains the keratin 5 promotor. Interestingly, the volume of eccrine sweat glands was reduced markedly in the K5-Brn2 transgenic mouse compared with the wild-type, while the expression of aquaporin 5, important molecule in sweat secretion, was increased in each sweat gland cell, probably to compensate for the reduction in gland development. However, sweating response to a pilocarpine injection in the hind paw was significantly decreased in the K5-Brn2 transgenic mouse compared with the wild-type. The paw epidermis was thicker in the K5-Brn2 transgenic mouse compared with the wild-type. Taken together, eccrine sweat gland development and sweat secretion were suppressed markedly in the K5-Brn2 transgenic mouse. These results may be associated with dominant development of the epidermis by Brn2 overexpression in the paw skin.

NFIB Mediates BRN2 Driven Melanoma Cell Migration and Invasion Through Regulation of EZH2 and MITF.

While invasion and metastasis of tumour cells are the principle factor responsible for cancer related deaths, the mechanisms governing the process remain poorly defined. Moreover, phenotypic divergence of sub-populations of tumour cells is known to underpin alternative behaviors linked to tumour progression such as proliferation, survival and invasion. In the context of melanoma, heterogeneity between two transcription factors, BRN2 and MITF, has been associated with phenotypic switching between predominantly invasive and proliferative behaviors respectively. Epigenetic changes, in response to external cues, have been proposed to underpin this process, however the mechanism by which the phenotypic switch occurs is unclear. Here we report the identification of the NFIB transcription factor as a novel downstream effector of BRN2 function in melanoma cells linked to the migratory and invasive characteristics of these cells. Furthermore, the function of NFIB appears to drive an invasive phenotype through an epigenetic mechanism achieved via the upregulation of the polycomb group protein EZH2. A notable target of NFIB mediated up-regulation of EZH2 is decreased MITF expression, which further promotes a less proliferative, more invasive phenotype. Together our data reveal that NFIB has the ability to promote dynamic changes in the chromatin state of melanoma cells to facilitate migration, invasion and metastasis.

Inhibition of oncogenic BRAF activity by indole-3-carbinol disrupts microphthalmia-associated transcription factor expression and arrests melanoma cell proliferation.

Indole-3-carbinol (I3C), an anti-cancer phytochemical derived from cruciferous vegetables, strongly inhibited proliferation and down-regulated protein levels of the melanocyte master regulator micropthalmia-associated transcription factor (MITF-M) in oncogenic BRAF-V600E expressing melanoma cells in culture as well as in vivo in tumor xenografted athymic nude mice. In contrast, wild type BRAF-expressing melanoma cells remained relatively insensitive to I3C anti-proliferative signaling. In BRAF-V600E-expressing melanoma cells, I3C treatment inhibited phosphorylation of MEK and ERK/MAPK, the down stream effectors of BRAF. The I3C anti-proliferative arrest was concomitant with the down-regulation of MITF-M transcripts and promoter activity, loss of endogenous BRN-2 binding to the MITF-M promoter, and was strongly attenuated by expression of exogenous MITF-M. Importantly, in vitro kinase assays using immunoprecipitated BRAF-V600E and wild type BRAF demonstrated that I3C selectively inhibited the enzymatic activity of the oncogenic BRAF-V600E but not of the wild type protein. In silico modeling predicted an I3C interaction site in the BRAF-V600E protomer distinct from where the clinically used BRAF-V600E inhibitor Vemurafenib binds to BRAF-V600E. Consistent with this prediction, combinations of I3C and Vemurafenib more potently inhibited melanoma cell proliferation and reduced MITF-M levels in BRAF-V600E expressing melanoma cells compared to the effects of each compound alone. Thus, our results demonstrate that oncogenic BRAF-V600E is a new cellular target of I3C that implicate this indolecarbinol compound as a potential candidate for novel single or combination therapies for melanoma. © 2016 Wiley Periodicals, Inc.

In silico studies of the interaction between BRN2 protein and MORE DNA.

The incidence of skin cancer has increased in recent decades, and melanoma is the most aggressive form with the lowest chance of successful treatment. Currently, drug design projects are in progress, but available treatments against metastatic melanoma have not significantly increased survival, and few patients are cured. Thus, new therapeutic agents should be developed as more effective therapeutic options for melanoma. High levels of the BRN2 transcription factor have been related to melanoma development. However, neither the three-dimensional (3D) structure of BRN2 protein nor its POU domain has been determined experimentally. Construction of the BRN2 3D structure, and the study of its interaction with its DNA target, are important strategies for increasing the structural and functional knowledge of this protein. Thus, the aim of this work was to study the interaction between BRN2 and MORE DNA through in silico methods. The full-length BRN2 3D structure was built using the PHYRE2 and Swiss-Model programs, and molecular dynamics of this protein in complex with MORE DNA was simulated for 20 ns by the NAMD program. The BRN2 model obtained includes helix and loop regions, and the BRN2 POU domain shares structural similarity with other members of the transcription factor family. No significant conformational change of this protein occurred during dynamics simulation. These analyses revealed BRN2 residues important for the specific interaction with nucleotide bases and with more than one DNA nucleotide. This study may contribute to the design of inhibitors against BRN2 or MORE DNA as molecular targets of melanoma skin cancer. Graphical Abstract Model of complete Brn2 protein in complex with MORE DNA after building through comparative modeling and refinement by molecular dynamics simulation.