The moon's monthly cycle synchronizes reproduction in countless marine organisms. The mass-spawning bristle worm Platynereis dumerilii uses an endogenous monthly oscillator set by full moon to phase ...reproduction to specific days. But how do organisms recognize specific moon phases? We uncover that the light receptor L-Cryptochrome (L-Cry) discriminates between different moonlight durations, as well as between sun- and moonlight. A biochemical characterization of purified L-Cry protein, exposed to naturalistic sun- or moonlight, reveals the formation of distinct sun- and moonlight states characterized by different photoreduction- and recovery kinetics of L-Cry's co-factor Flavin Adenine Dinucleotide. In Platynereis, L-Cry's sun- versus moonlight states correlate with distinct subcellular localizations, indicating different signaling. In contrast, r-Opsin1, the most abundant ocular opsin, is not required for monthly oscillator entrainment. Our work reveals a photo-ecological concept for natural light interpretation involving a "valence interpreter" that provides entraining photoreceptor(s) with light source and moon phase information.
Organisms generate shapes across size scales. Whereas patterning and morphogenesis of macroscopic tissues has been extensively studied, the principles underlying the formation of micrometric and ...submicrometric structures remain largely enigmatic. Individual cells of polychaete annelids, so-called chaetoblasts, are associated with the generation of chitinous bristles of highly stereotypic geometry. Here we show that bristle formation requires a chitin-producing enzyme specifically expressed in the chaetoblasts. Chaetoblasts exhibit dynamic cell surfaces with stereotypical patterns of actin-rich microvilli. These microvilli can be matched with internal and external structures of bristles reconstructed from serial block-face electron micrographs. Individual chitin teeth are deposited by microvilli in an extension-disassembly cycle resembling a biological 3D printer. Consistently, pharmacological interference with actin dynamics leads to defects in tooth formation. Our study reveals that both material and shape of bristles are encoded by the same cell, and that microvilli play a role in micro- to submicrometric sculpting of biomaterials.
Significance The moon provides highly reliable time information to organisms. Whereas sunlight is known to set daily animal timing systems, mechanistic insight into the impact of moonlight on such ...systems remains scarce. We establish that the marine bristleworm
times the precise hours of mass spawning by integrating lunar light information into a plastic daily timing system able to run with circadian (∼24 h) or circalunidian (∼24.8 h) periodicity. The correct interpretation of moonlight is mediated by the interplay of two light sensors: a cryptochrome and a melanopsin ortholog provide information on light valence and moonrise time, respectively. Besides its ecological relevance, our work provides a plausible explanation for long-standing observations of light intensity-dependent differences in circadian clock periods.
Rhabdomeric opsins (r-opsins) are light sensors in cephalic eye photoreceptors, but also function in additional sensory organs. This has prompted questions on the evolutionary relationship of these ...cell types, and if ancient r-opsins were non-photosensory. A molecular profiling approach in the marine bristleworm
revealed shared and distinct features of cephalic and non-cephalic
-expressing cells. Non-cephalic cells possess a full set of phototransduction components, but also a mechanosensory signature. Prompted by the latter, we investigated
putative mechanotransducer and found that
and
co-expressed with
in TRE cells by HCR RNA-FISH. To further assess the role of r-Opsin1 in these cells, we studied its signaling properties and unraveled that r-Opsin1 is a Gαq-coupled blue light receptor. Profiling of cells from
mutants versus wild-types, and a comparison under different light conditions reveals that in the non-cephalic cells light - mediated by r-Opsin1 - adjusts the expression level of a calcium transporter relevant for auditory mechanosensation in vertebrates. We establish a deep-learning-based quantitative behavioral analysis for animal trunk movements and identify a light- and r-Opsin-1-dependent fine-tuning of the worm's undulatory movements in headless trunks, which are known to require mechanosensory feedback. Our results provide new data on peripheral cell types of likely light sensory/mechanosensory nature. These results point towards a concept in which such a multisensory cell type evolved to allow for fine-tuning of mechanosensation by light. This implies that light-independent mechanosensory roles of r-opsins may have evolved secondarily.
Ubiquitinated proteins can alternatively be delivered directly to the proteasome or via p97/VCP (valosin-containing protein). Whereas the proteasome degrades ubiquitinated proteins, the homohexameric ...ATPase p97/VCP seems to control the ubiquitination status of recruited substrates. The COP9 signalosome (CSN) is also involved in the ubiquitin/proteasome system (UPS) as exemplified by regulating the neddylation of ubiquitin E3 ligases. Here, we show that p97/VCP colocalizes and directly interacts with subunit 5 of the CSN (CSN5) in vivo and is associated with the entire CSN complex in an ATP-dependent manner. Furthermore, we provide evidence that the CSN and in particular the isopeptidase activity of its subunit CSN5 as well as the associated deubiquitinase USP15 are required for proper processing of polyubiquitinated substrates bound to p97/VCP. Moreover, we show that in addition to NEDD8, CSN5 binds to oligoubiquitin chains in vitro. Therefore, CSN and p97/VCP could form an ATP-dependent complex that resembles the 19 S proteasome regulatory particle and serves as a key mediator between ubiquitination and degradation pathways.
We explored the mechanisms leading to the distinct overexpression of EPOR as well as the effects of EPO signaling on ETV6/RUNX1-positive acute lymphoblastic leukemias.
ETV6/RUNX1-expressing model ...cell lines and leukemic cells were used for real-time PCR of EPOR expression. Proliferation, viability, and apoptosis were analyzed on cells exposed to EPO, prednisone, or inhibitors of EPOR pathways by 3Hthymidine incorporation, 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay, and Annexin V/propidium iodide staining. Western blot analysis was done to detect activation of signaling proteins. Serum EPO levels and sequences of the EPOR (n = 53) as well as hemoglobin levels were taken from children with acute lymphoblastic leukemia enrolled in Austrian protocols.
We show here that ectopic expression of ETV6/RUNX1 induced EPOR up-regulation. Anemia, however, did not appear to influence EPOR expression on leukemic cells, although children with ETV6/RUNX1-positive leukemias had a lower median hemoglobin than controls. Exposure to EPO increased proliferation and survival of ETV6/RUNX1-positive leukemias in vitro, whereas blocking its binding site did not alter cell survival. The latter was not caused by activating mutations in the EPOR but might be triggered by constitutive activation of phosphatidylinositol 3-kinase/Akt, the major signaling pathway of EPOR in these cells. Moreover, prednisone-induced apoptosis was attenuated in the presence of EPO in this genetic subgroup.
Our data suggest that ETV6/RUNX1 leads to EPOR up-regulation and that activation by EPO might be of relevance to the biology of this leukemia subtype. Further studies are, however, needed to assess the clinical implications of its apoptosis-modulating properties.
Mitochondria are fundamental for life and require balanced ion exchange to maintain proper functioning. The mitochondrial cation exchanger LETM1 sparks interest because of its pathophysiological role ...in seizures in the Wolf Hirschhorn Syndrome (WHS). Despite observation of sleep disorganization in epileptic WHS patients, and growing studies linking mitochondria and epilepsy to circadian rhythms, LETM1 has not been studied from the chronobiological perspective. Here we established a viable
knock-out, using the diurnal vertebrate
to study the metabolic and chronobiological consequences of
deficiency. We report diurnal rhythms of Letm1 protein levels in wild-type fish. We show that mitochondrial nucleotide metabolism is deregulated in
mutant fish, the rate-limiting enzyme of NAD
production is up-regulated, while NAD
and NADH pools are reduced. These changes were associated with increased expression amplitude of circadian core clock genes in
compared with wild-type under light/dark conditions, suggesting decreased NAD(H) levels as a possible mechanism for circadian system perturbation in Letm1 deficiency. Replenishing NAD pool may ameliorate WHS-associated sleep and neurological disorders.
Metastasis is associated with poor prognosis for melanoma responsible for about 90% of skin cancer‐related mortality. To metastasize, melanoma cells must escape keratinocyte control, invade across ...the basement membrane and survive in the dermis by resisting apoptosis before they can intravasate into the circulation. α‐Catulin (CTNNAL1) is a cytoplasmic molecule that integrates the crosstalk between nuclear factor‐kappa B and Rho signaling pathways, binds to β‐catenin and increases the level of both α‐catenin and β‐catenin and therefore has potential effects on inflammation, apoptosis and cytoskeletal reorganization. Here, we show that α‐catulin is highly expressed in melanoma cells. Expression of α‐catulin promoted melanoma progression and occurred concomitantly with the downregulation of E‐cadherin and the upregulation of expression of mesenchymal genes such as N‐cadherin, Snail/Slug and the matrix metalloproteinases 2 and 9. Knockdown of α‐catulin promoted adhesion to and inhibited migration away from keratinocytes in an E‐cadherin‐dependent manner and decreased the transmigration through a keratinocyte monolayer, as well as in Transwell assays using collagens, laminin and fibronectin coating. Moreover, knockdown promoted homotypic spheroid formation and concomitantly increased E‐cadherin expression along with downregulation of transcription factors implicated in its repression (Snail/Slug, Twist and ZEB). Consistent with the molecular changes, α‐catulin provoked invasion of melanoma cells in a three‐dimensional culture assay by the upregulation of matrix metalloproteinases 2 and 9 and the activation of ROCK/Rho. As such, α‐catulin may represent a key driver of the metastatic process, implicating potential for therapeutic interference.
What's new?
The authors showed for the first time that alpha‐catulin is highly expressed in melanoma cells. alpha‐catulin knockdown altered the expression of E‐cadherin and other genes known to be implicated in melanoma progression. Furthermore, knockdown of alpha‐catulin promoted both binding of melanoma cells to keratinocytes and spheroid formation by enhanced E‐cadherin expression. The authors also found that alpha‐catulin provoked invasion of melanoma cells in a 3D culture assay by up‐regulation of the matrix metalloproteinases 2 and 9 and activation of Rho. alpha‐catulin may thus represent a key driver of the metastatic process in human melanoma, implicating potential for therapeutic interference.
Metastasis is associated with poor prognosis for melanoma responsible for about 90% of skin cancer-related mortality. To metastasize, melanoma cells must escape keratinocyte control, invade across ...the basement membrane and survive in the dermis by resisting apoptosis before they can intravasate into the circulation. alpha-Catulin (CTNNAL1) is a cytoplasmic molecule that integrates the crosstalk between nuclear factor-kappa B and Rho signaling pathways, binds to beta-catenin and increases the level of both alpha-catenin and beta-catenin and therefore has potential effects on inflammation, apoptosis and cytoskeletal reorganization. Here, we show that alpha-catulin is highly expressed in melanoma cells. Expression of alpha-catulin promoted melanoma progression and occurred concomitantly with the downregulation of E-cadherin and the upregulation of expression of mesenchymal genes such as N-cadherin, Snail/Slug and the matrix metalloproteinases 2 and 9. Knockdown of alpha-catulin promoted adhesion to and inhibited migration away from keratinocytes in an E-cadherin-dependent manner and decreased the transmigration through a keratinocyte monolayer, as well as in Transwell assays using collagens, laminin and fibronectin coating. Moreover, knockdown promoted homotypic spheroid formation and concomitantly increased E-cadherin expression along with downregulation of transcription factors implicated in its repression (Snail/Slug, Twist and ZEB). Consistent with the molecular changes, alpha-catulin provoked invasion of melanoma cells in a three-dimensional culture assay by the upregulation of matrix metalloproteinases 2 and 9 and the activation of ROCK/Rho. As such, alpha-catulin may represent a key driver of the metastatic process, implicating potential for therapeutic interference. What's new? The authors showed for the first time that alpha-catulin is highly expressed in melanoma cells. alpha-catulin knockdown altered the expression of E-cadherin and other genes known to be implicated in melanoma progression. Furthermore, knockdown of alpha-catulin promoted both binding of melanoma cells to keratinocytes and spheroid formation by enhanced E-cadherin expression. The authors also found that alpha-catulin provoked invasion of melanoma cells in a 3D culture assay by up-regulation of the matrix metalloproteinases 2 and 9 and activation of Rho. alpha-catulin may thus represent a key driver of the metastatic process in human melanoma, implicating potential for therapeutic interference. PUBLICATION ABSTRACT
The stress inducible heat shock protein 70 (hsp70) confers resistance to a variety of adverse environmental conditions including certain anticancer drugs. In the present study we explored cellular ...consequences of hsp70 overexpression with respect to genotoxic stress. Employing an isogenic set of stable transfectant cells, one overexpressing hsp 70 and the other serving as control, we found that a high hsp70 expression level is sufficient to provide protection against the cytotoxicity of doxorubicin. In addition, hsp70-protected cells showed the capability to restart cell proliferation at concentrations where control cells arrested. However, the DNA lesion density was comparable in the lines used. Recording the cell cycle control revealed a dramatic shortening of the doxorubicin-mediated cell cycle arrest in the G2 phase upon hsp70 overexpression. Our data suggest an involvement of hsp70 in the regulation of the cell cycle.
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