RESUMO
Genome instability relies on preserving the chromatin structure, with any histone imbalances threating DNA integrity. Histone synthesis occurs in the cytoplasm, followed by a maturation process before their nuclear translocation. This maturation involves protein folding and the establishment of post-translational modifications. Disruptions in this pathway hinder chromatin assembly and contribute to genome instability. JMJD1B, a histone demethylase, not only regulates gene expression but also ensures a proper supply of histones H3 and H4 for the chromatin assembly. Reduced JMJD1B levels lead to the cytoplasmic accumulation of histones, causing defects in the chromatin assembly and resulting in DNA damage. To investigate the role of JMJD1B in regulating genome stability and the malignancy of melanoma tumors, we used a JMJD1B/KDM3B knockout in B16F10 mouse melanoma cells to perform tumorigenic and genome instability assays. Additionally, we analyzed the transcriptomic data of human cutaneous melanoma tumors. Our results show the enhanced tumorigenic properties of JMJD1B knockout melanoma cells both in vitro and in vivo. The γH2AX staining, Micrococcal Nuclease sensitivity, and comet assays demonstrated increased DNA damage and genome instability. The JMJD1B expression in human melanoma tumors correlates with a lower mutational burden and fewer oncogenic driver mutations. Our findings highlight JMJD1B's role in maintaining genome integrity by ensuring a proper histone supply to the nucleus, expanding its function beyond gene expression regulation. JMJD1B emerges as a crucial player in preserving genome stability and the development of melanoma, with a potential role as a safeguard against oncogenic mutations.
Assuntos
Dano ao DNA , Instabilidade Genômica , Histonas , Histona Desmetilases com o Domínio Jumonji , Melanoma , Neoplasias Cutâneas , Animais , Humanos , Camundongos , Linhagem Celular Tumoral , Dano ao DNA/genética , Regulação Neoplásica da Expressão Gênica , Histonas/metabolismo , Histona Desmetilases com o Domínio Jumonji/metabolismo , Histona Desmetilases com o Domínio Jumonji/genética , Melanoma/genética , Melanoma/patologia , Melanoma/metabolismo , Melanoma Experimental/genética , Melanoma Experimental/patologia , Melanoma Experimental/metabolismo , Neoplasias Cutâneas/genética , Neoplasias Cutâneas/patologia , Neoplasias Cutâneas/metabolismoRESUMO
Transcription factors are multidomain proteins with specific DNA binding and regulatory domains. In the human FoxP subfamily (FoxP1, FoxP2, FoxP3, and FoxP4) of transcription factors, a 90 residue-long disordered region links a Leucine Zipper (ZIP)-known to form coiled-coil dimers-and a Forkhead (FKH) domain-known to form domain swapping dimers. We used replica exchange discrete molecular dynamics simulations, single-molecule fluorescence experiments, and other biophysical tools to understand how domain tethering in FoxP1 impacts dimerization at ZIP and FKH domains and how DNA binding allosterically regulates their dimerization. We found that domain tethering promotes FoxP1 dimerization but inhibits a FKH domain-swapped structure. Furthermore, our findings indicate that the linker mediates the mutual organization and dynamics of ZIP and FKH domains, forming closed and open states with and without interdomain contacts, thus highlighting the role of the linkers in multidomain proteins. Finally, we found that DNA allosterically promotes structural changes that decrease the dimerization propensity of FoxP1. We postulate that, upon DNA binding, the interdomain linker plays a crucial role in the gene regulatory function of FoxP1.
Assuntos
DNA , Fatores de Transcrição , Humanos , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Dimerização , DNA/química , Domínios Proteicos , Regulação da Expressão Gênica , Proteínas Repressoras/química , Proteínas Repressoras/genética , Proteínas Repressoras/metabolismo , Fatores de Transcrição Forkhead/química , Fatores de Transcrição Forkhead/genética , Fatores de Transcrição Forkhead/metabolismoRESUMO
The Plasmodium falciparum multigene var family codes for approximately 50 variant adhesive proteins expressed in a mutually exclusive manner at the surface of infected red blood cells (iRBCs). Switching expression of var genes can lead to fundamental changes in the adhesive and antigenic properties of iRBCs. For example, a specific phenotypic switch in adhesion from CD36 to chondroitin sulphate A (CSA) is associated with malaria pathogenesis in pregnant women. The factors and DNA elements that control the expression of a particular member of the var gene family during gestational malaria remains enigmatic. Here, we report that the subtelomeric FCR3 varCSA is expressed under the control of a unique DNA element of 1.8 kb, whereas the other members of the var multigene family are flanked by common regulatory elements. The 5' varCSA-type element is conserved as a single copy in laboratory strains and clinical isolates from Brazil and West Africa and contains two distinct repetitive elements of 150 bp and 60 bp respectively. The 5' varCSA-type sequence tags a var gene in the 3D7 genome that is homologous to the FCR3 varCSA gene. A recombinant DBL gamma domain of this var gene showed specific binding to CSA. This subtelomeric varCSA gene is transcribed in the opposite sense when compared with the usual orientation of telomere-adjacent var genes. This unique arrangement might explain why the varCSA gene is relatively conserved in genetically distinct parasites despite being located in a highly recombinogenic chromosome compartment. The 5' untranslated region (UTR) of the varCSA-type sequence is also transcribed in placental isolates that bind to CSA, illustrating an important role for the unique 5' varCSA-type sequence in the regulation of var genes involved in malaria pathogenesis in pregnant women. However, this promoter is not always found to be transcribing var genes selected for expression of products that bind to CSA in vitro. Our work identifies a sequence tag for the identification of varCSA genes in placental isolates for the first time.