RESUMO
Programmed cell death (PCD) is a common cell fate in metazoan development. PCD effectors are extensively studied, but how they are temporally regulated is less understood. Here, we report a mechanism controlling tail-spike cell death onset during Caenorhabditis elegans development. We show that the zinc-finger transcription factor BLMP-1, which controls larval development timing, also regulates embryonic tail-spike cell death initiation. BLMP-1 functions upstream of CED-9 and in parallel to DRE-1, another CED-9 and tail-spike cell death regulator. BLMP-1 expression is detected in the tail-spike cell shortly after the cell is born, and blmp-1 mutations promote ced-9-dependent tail-spike cell survival. BLMP-1 binds ced-9 gene regulatory sequences, and inhibits ced-9 transcription just before cell-death onset. BLMP-1 and DRE-1 function together to regulate developmental timing, and their mammalian homologs regulate B-lymphocyte fate. Our results, therefore, identify roles for developmental timing genes in cell-death initiation, and suggest conservation of these functions.
Assuntos
Proteínas de Caenorhabditis elegans/genética , Caenorhabditis elegans/genética , Morte Celular/genética , Proteínas Repressoras/genética , Transcrição Gênica/genética , Animais , Apoptose/genética , Diferenciação Celular/genética , Regulação da Expressão Gênica no Desenvolvimento/genéticaRESUMO
Skin epidermis secretes apical extracellular matrix (aECM) as a protective barrier from the external environment. The aECM is highly dynamic and constantly undergoes remodeling during animal development. How aECM dynamics is temporally regulated during development, and whether and how its mis-regulation may impact epidermal cell morphology or function remains to be fully elucidated. Here, we report that the conserved Zn-finger transcription factor BLMP-1/Blimp1, which regulates epidermal development in C. elegans, controls apical cell shape of the epidermis by downregulation of aECM remodeling. Loss of blmp-1 causes upregulation of genes essential for molting, including bus-8 and mlt-8, in adult, leading to an abnormal shape in the apical region of adult epidermal cells. The apical epidermal morphological defect is suppressed by reduction of bus-8 or mlt-8. BUS-8 is a key mannosyltransferase, which functions in glycosylation of N-linked glycoproteins; MLT-8 has a ganglioside GM2 lipid-binding domain and is implicated in signaling during molting, a process where the old cuticle is shed and synthesized anew. Overexpression of bus-8 or mlt-8 induces an apical epidermal cell defect as observed in blmp-1 mutants. MLT-8::GFP fusion protein is localized to lysosomes and secreted to aECM. BUS-8 is important for MLT-8 stability and lysosomal targeting, which may be regulated by BUS-8-mediated glycosylation of MLT-8 and function as a molting signaling cue in aECM remodeling. We propose that BLMP-1 represses MLT-8 expression and glycosylation in the epidermis to prevent inappropriate aECM remodeling, which is essential for maintenance of apical epidermal cell morphology during larva-to-adult transition.
Assuntos
Proteínas de Caenorhabditis elegans , Caenorhabditis elegans , Animais , Caenorhabditis elegans/metabolismo , Proteínas de Caenorhabditis elegans/genética , Proteínas de Caenorhabditis elegans/metabolismo , Células Epidérmicas/metabolismo , Epiderme/metabolismo , Manosiltransferases/metabolismo , Muda/genéticaRESUMO
Matching the treatment to an individual patient's tumor state can increase therapeutic efficacy and reduce tumor recurrence. Circulating tumor cells (CTCs) derived from solid tumors are promising subjects for theragnostic analysis. To analyze how CTCs represent tumor states, we established cell lines from CTCs, primary and metastatic tumors from a mouse model and provided phenotypic and multiomic analyses of these cells. CTCs and metastatic cells, but not primary tumor cells, shared stochastic mutations and similar hypomethylation levels at transcription start sites. CTCs and metastatic tumor cells shared a hybrid epithelial/mesenchymal transcriptome state with reduced adhesive and enhanced mobilization characteristics. We tested anti-cancer drugs on tumor cells from a metastatic breast cancer patient. CTC responses mirrored the impact of drugs on metastatic rather than primary tumors. Our multiomic and clinical anti-cancer drug response results reveal that CTCs resemble metastatic tumors and establish CTCs as an ex vivo tool for personalized medicine.
RESUMO
More than 55 million people live with dementia worldwide in 2021, and there are nearly 10 million new cases every year. Alzheimer's disease (AD) is the most common cause of dementia. Despite urgent need, early detection of AD and long-term monitoring of AD progression have been challenging. This is due to the limited availability of brain imaging facilities and the highly invasive procedure with the cerebrospinal fluid assay to assess the level of AD biomarkers, such as beta-amyloid (Aß). Reliable measurements of AD biomarkers in blood samples are still difficult because of their very low abundance. Here, we develop a rapid, specific, and ultrasensitive immunoassay using plasmonic-gold nanoisland (pGOLD) chips with near-infrared fluorescence-enhanced detection for Aß1-40 and Aß1-42. We show step-by-step processes and results during the platform establishment, including antibody specificity and sensitivity tests, antibody pair examination, condition optimization, and procedure refinement. Finally, we demonstrate the platform performance with detection sensitivity at the subpicogram per milliliter level. This platform, therefore, has a great application potential for early detection of AD using blood samples.
Assuntos
Doença de Alzheimer/diagnóstico , Peptídeos beta-Amiloides/análise , Materiais Biomiméticos/química , Ouro/química , Nanopartículas Metálicas/química , Doença de Alzheimer/sangue , Humanos , Imunoensaio , Teste de Materiais , Tamanho da PartículaRESUMO
Peptide neuromodulators are released from a unique organelle: the dense-core vesicle. Dense-core vesicles are generated at the trans-Golgi and then sort cargo during maturation before being secreted. To identify proteins that act in this pathway, we performed a genetic screen in Caenorhabditis elegans for mutants defective in dense-core vesicle function. We identified two conserved Rab2-binding proteins: RUND-1, a RUN domain protein, and CCCP-1, a coiled-coil protein. RUND-1 and CCCP-1 colocalize with RAB-2 at the Golgi, and rab-2, rund-1, and cccp-1 mutants have similar defects in sorting soluble and transmembrane dense-core vesicle cargos. RUND-1 also interacts with the Rab2 GAP protein TBC-8 and the BAR domain protein RIC-19, a RAB-2 effector. In summary, a pathway of conserved proteins controls the maturation of dense-core vesicles at the trans-Golgi network.
Assuntos
Proteínas de Caenorhabditis elegans/metabolismo , Mutação/genética , Vesículas Secretórias/metabolismo , Proteína rab2 de Ligação ao GTP/metabolismo , Análise de Variância , Animais , Animais Geneticamente Modificados , Caenorhabditis elegans , Proteínas de Caenorhabditis elegans/genética , Mapeamento Cromossômico , Clonagem Molecular , Endossomos/metabolismo , Endossomos/ultraestrutura , Complexo de Golgi/metabolismo , Complexo de Golgi/ultraestrutura , Proteínas de Fluorescência Verde/genética , Proteínas de Fluorescência Verde/metabolismo , Microscopia Eletrônica de Transmissão , Dados de Sequência Molecular , Transporte Proteico , Vesículas Secretórias/genética , Vesículas Secretórias/ultraestrutura , Proteína rab2 de Ligação ao GTP/genéticaRESUMO
Mutations that affect the Z-disk-associated ALP-Enigma proteins have been linked to human muscular and cardiac diseases. Despite their clear physiological significance for human health, the mechanism of action of ALP-Enigma proteins is largely unknown. In Caenorhabditis elegans, the ALP-Enigma protein family is encoded by a single gene, alp-1; thus C. elegans provides an excellent model to study ALP-Enigma function. Here we present a molecular and genetic analysis of ALP-Enigma function in C. elegans. We show that ALP-1 and alpha-actinin colocalize at dense bodies where actin filaments are anchored and that the proper localization of ALP-1 at dense bodies is dependent on alpha-actinin. Our analysis of alp-1 mutants demonstrates that ALP-1 functions to maintain actin filament organization and participates in muscle stabilization during contraction. Reducing alpha-actinin activity enhances the actin filament phenotype of the alp-1 mutants, suggesting that ALP-1 and alpha-actinin function in the same cellular process. Like alpha-actinin, alp-1 also interacts genetically with a connectin/titin family member, ketn-1, to provide mechanical stability for supporting body wall muscle contraction. Taken together, our data demonstrate that ALP-1 and alpha-actinin function together to stabilize actin filaments and promote muscle structural integrity.
Assuntos
Citoesqueleto de Actina/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Proteínas de Caenorhabditis elegans/metabolismo , Caenorhabditis elegans/metabolismo , Músculos/metabolismo , Actinina/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/genética , Animais , Especificidade de Anticorpos , Fenômenos Biomecânicos , Caenorhabditis elegans/citologia , Proteínas de Caenorhabditis elegans/genética , Estruturas Celulares/metabolismo , Conectina , Proteínas com Domínio LIM , Proteínas dos Microfilamentos/metabolismo , Proteínas Musculares/metabolismo , Músculos/citologia , Proteínas Mutantes/metabolismo , Mutação/genética , Transporte Proteico , Vinculina/metabolismoRESUMO
Members of the ALP/Enigma family of PDZ-LIM proteins play a role in cytoskeletal anchorage and mutations in at least one member of this family are associated with human cardiomyopathy. Here, we describe the analysis of the Caenorhabditis elegans alp-1 gene. alp-1 is predicted to encode the entire nematode ALP/Enigma protein family, consisting of one ALP-related protein with a single LIM domain and three Enigma-like proteins containing four LIM domains. We demonstrate that the ALP-1 proteins are expressed in muscle cells, where they localize to actin anchorage and muscle attachment sites. We show that the PDZ domain of the ALP-1 proteins is sufficient to target the protein to the dense bodies, which are important actin anchorage sites in C. elegans body wall muscle. We demonstrate that the C. elegans ALP/Enigma proteins are also localized to cell-cell junctions and to both epithelial and muscle cell nuclei. These findings suggest new roles for the ALP/Enigma protein family that may lead to the understanding of their involvement in cardiomyopathy.