Biochemistry (Moscow)最新文献

Guanylins are intestinal natriuretic peptides that regulate water and electrolyte balance in the intestine and kidney. Their primary receptor is membrane guanylyl cyclase C (GC-C), while alternative functions related to feeding behavior and olfactory preferences are mediated by guanylyl cyclase D (GC-D) expressed exclusively in olfactory neurons. Evidence suggests existence of unidentified receptors activated by guanylin peptides in the absence of GC-C that affect sodium metabolism. Some of these receptors trigger the cGMP-dependent signaling pathways typical exclusively for guanylyl cyclases. This review provides a comparative analysis of the existing data on different membrane receptor guanylyl cyclases, including early discoveries and contemporary research, focusing on their potential as guanylin targets.

Reviewing published concepts on the chemical interactions between small molecules implicated in the origin of life suggests that their chemical properties have included not only those that might have been suitable for metabolic pathways. Some of the immanent “excessive” potencies of molecules make them able to form covalent adducts with proteins and nucleic acids. The accumulation of macromolecules damaged in this way could decrease the viability of protocells with increasing age. Thus, aging (senescence) could emerge concomitantly with life as its chemical heritage. Moreover, the exponential increase in mortality with age (the Gompertz law) could emerge when the kinetics of molecular disintegration according to the Arrhenius equation (disintegration rate depends exponentially on varying temperature at a constant activation barrier) was inherited by the kinetics of protocells dying out in their populations, the role of the independent variable passing from temperature, which was virtually constant on the Calvin scale, to viability. The cooperation of these two chemical heritages was enough to eliminate effectively old living objects and to make any evolved program of aging needless. Therefore, aging had not resulted from the biological evolution but rather has been and still is its independent factor. All this was possible without oxygen, which could only modify, rather than form de novo, the primary chemical driving force of aging. With all that, the energy benefits of aerobic metabolism have provided for the advent of multicellular organisms, in particular, those featuring massive extracellular matter and unrenewable cell populations, including those comprising the brain. Their functions are incompatible with complete renewal. This makes the role of oxygen in aging not limited to being the source of reactive oxygen species. Oxygen had been indispensable for the advent of both accumulators of chemical damage and ability to recognize it. In a sense, it was not a problem for nature to develop aging in the course of evolution towards humans, for whom being aware of aging is a problem. Its satisfactory solution cannot be chemical, physical, pharmacological, or otherwise technical. It can only be mental.

Skeletal muscle unloading results in muscle atrophy associated with the upregulation of proteolytic genes and suppression of protein synthesis, often accompanied by altered calcium signaling. Here, we used the inositol trisphosphate receptor (IP3R) inhibitor aminoethoxydiphenyl borate (2-APB) to explore the hypothesis that these changes are mediated by IP3Rs. Male Wistar rats were divided into 4 groups: (i) control, (ii) control with daily injections of 2-APB, (iii) 3 days of hind limb suspension, (iv) 3 days of hind limb suspension with daily administration of 2-APB. At the end-point, soleus muscles from the animals were analyzed by Western blotting for the markers of calcium, anabolic, and catabolic signaling. The 3-day hind limb unloading resulted in a decreased muscle weight index, upregulation of the anabolic suppressor pThr56-eEF2, downregulation of anabolic signaling via the mTOR pathway and rRNA expression, as well as the increase in the content of nuclear pThr286-CaMKII (p < 0.05) and cytosolic calcineurin A. While 2-APB did not affect the mTOR-governed changes in anabolism and catabolism, it significantly attenuated alterations in the calcium-dependent targets, such as CaMKII, calcineurin, and eEF2. By contrast, proteolytic signaling (expression of MuRF1, atrogin-1, Ulk1, and ubiquitin mRNAs) after 3-day hind limb unloading was equally upregulated in the control and 2-APB-treated animals. These results suggest that IP3Rs are involved in the unloading-induced muscle atrophy by controlling the nuclear content of calcium; however, they are dispensable for reduced mTOR activity and altered metabolism.

In euryhaline fish species, including the three-spined stickleback, a key physiological response to freshwater adaptation aimed at maintaining osmotic homeostasis is enhancement of ion uptake from the environment and reduction of ion loss. Hormone prolactin, a central regulator of this process, primarily targets gills and intestine. Our previous work demonstrated that in the model of freshwater adaptation in sticklebacks prolactin expression and sensitivity of osmoregulatory tissues to prolactin differ between the males and females. In the present study, we measured expression levels of the genes encoding α1a and α3a subunits of Na⁺/K⁺-ATPase, as well as ion transporters NKCC1a, NKCC2, NCC, and NHE2, in the gill and intestinal tissues of the male and female three-spined sticklebacks (Gasterosteus aculeatus L.) under conditions of acute (24 h) and chronic (72 h) freshwater adaptation, relative to the control conditions. During the freshwater adaptation, females, but not males, exhibited increased intestinal expression of nhe2 and atp1a3 genes (as well as of the ratio of atp1a1/atp1a3 expression), and the nkcc1a gene, along with the decreased expression of the nkcc2 gene. In contrast, only males showed increase in the ncc gene expression in the intestine. In both sexes, exposure to fresh water led to the significant decrease in the nkcc1a gene expression in the gills. These findings support our hypothesis of sex-dependent plasticity in osmoregulatory function in sticklebacks, with females exhibiting a more pronounced response. This pattern further aligns with the previously reported stronger activation of the prolactin axis in the females under freshwater adaptation conditions.

Actin cytoskeleton is a key participant in numerous cellular processes, including organelle transport, motility, contractility, exocytosis, and endocytosis. It also plays a critical role in pathological processes such as malignant cancer cell invasion. The actin-binding proteins, particularly tropomyosins (Tpm) and cofilins, are involved in actin cytoskeleton remodeling. For this study, we selected the least studied isoforms of Tpm expressed from the TPM1 gene – Tpm1.7, Tpm1.8, and Tpm1.9 – as well as the more well-known Tpm1.1 and Tpm1.6. We investigated mutual influence of these Tpm isoforms and cofilin-1 (cof-1) on actin filament dynamics. Using co-sedimentation assays, we demonstrated that Tpm1.7, Tpm1.8, and Tpm1.9 significantly inhibit cof-1 binding to the F-actin surface. Viscometry was employed to assess depolymerizing and severing effects of cof-1 on actin filaments. Tpm1.1, Tpm1.8, and Tpm1.6 effectively prevented depolymerizing/severing action of cof-1, while the protective effect of Tpm1.7 and Tpm1.9 was less pronounced. The rhodamine-phalloidin displacement assay was used to analyze the cof-1-induced conformational changes in F-actin. All studied Tpm isoforms effectively prevented effects of cof-1 on actin filaments. Our findings indicate that the TPM1 gene products generally exert an inhibitory effect on cof-1 activity in relation to actin filament polymerization/depolymerization dynamics. Such properties of Tpm isoforms could be important for formation of specific intracellular populations of actin filaments.

One of the hallmarks of malignant neoplasms is their ability to sustain growth under hypoxic conditions resulting from insufficient oxygenation of tumor tissues. Prolonged hypoxia is associated with the gradual adaptation of tumor cells to low oxygen levels, leading to the enhanced survival, increased metastatic potential, and development of resistance to anticancer therapies. The aim of this study was to investigate the mechanisms underlying adaptation of breast cancer cell to prolonged hypoxia and maintenance of the hypoxia-tolerant phenotype. Using long-term culturing under low oxygen conditions (1% O₂), we established hypoxia-adapted sublines of luminal (MCF-7) and triple-negative (MDA-MB-231) breast cancer cells, characterized by a stable growth in a hypoxic environment. The acquisition of hypoxia tolerance was accompanied by the activation of the HIF-1α-dependent transcription factor STAT3 and persistent overexpression of Snail, a key downstream effector of STAT3. The maintenance and stabilization of hypoxia-tolerant phenotype are mediated by miR-181a-2, which targets the STAT3/Snail signaling axis in the resistant cells. Analysis of DNA methylation status revealed no significant changes in the expression or activity of DNA methyltransferases (DNMTs) in the hypoxia-adapted cells. However, pharmacological inhibition of DNMTs using decitabine, as well as DNMT knockdown, increased cell sensitivity to hypoxia and partially reversed the hypoxia-resistant phenotype, which was accompanied by the activation of pro-apoptotic p53 signaling. In conclusion, our findings suggest that the acquired hypoxia tolerance in breast cancer cells is mediated, at least in part, by the activation of the miR-181a-2/STAT3/Snail signaling pathway. Furthermore, the use of demethylating agents may represent a promising therapeutic approach to targeting hypoxia-tolerant cancer cell populations.

Autophagy not only helps eliminate damaged, mutated, or genomically unstable cells, but also increases the chances of tumor cells overcoming the consequences of damage caused by chemotherapy. Autophagy induced by anthracyclines is cytoprotective in most tumor cell lines. Pharmacological or genetic blocking of autophagy in this case sensitizes tumor cells to therapy. Activation of cytoprotective autophagy can lead to chemoresistance, and with excessive enhancement, it can lead to energy depletion and trigger autophagic death. In some cases, cytotoxic autophagy develops under the action of anthracyclines, and its blocking increases cell survival. Activation of cytotoxic autophagy, on the contrary, triggers the process of “self-eating.” Modulation of autophagy in response to chemotherapeutic agents can be a double-edged sword for tumor cells, leading to both death and survival.

Melting of promoter DNA around the transcription start site (TSS) is a critical step of transcription required for initiation of RNA synthesis. In bacteria, promoter melting is mediated by the holoenzyme of RNA polymerase (RNAP) consisting of the catalytic core enzyme and the promoter recognition subunit, σ factor. Previously, we showed that RNAPs from thermophilic Thermus aquaticus and mesophilic Deinococcus radiodurans are unable to open promoters at ambient temperatures and require heating for DNA melting. These properties depend on their σ factors and are recapitulated in the hybrid holoenzymes including these σ factors and the core enzyme of Escherichia coli. Here, we show that DNA supercoiling alleviates the observed cold-sensitivity of promoter opening by the Deinococcus-Thermus RNAPs and by the hybrid holoenzymes and allows melting of the transcription start site at the same temperatures as in the case of E. coli RNAP. Supercoiling also suppresses salt sensitivity of the promoter complexes formed by these RNAPs. The results demonstrate that the RNAPs from Deinococcus-Thermus species are sensitive to DNA supercoiling and suggest that they can be rapidly switched-off or activated by the supercoiling state of the host genomes.

A growing body of evidence indicates a high incidence of osteoarthritis (OA) in both weight-bearing and non-weight-bearing joints in obese patients. The levels of leptin in the synovial fluid of obese patients with OA are elevated compared to healthy people, suggesting that leptin may be a key factor of OA in obese individuals. Synovitis can occur at all stages of OA development, causing diseases progression. We examined the effect of leptin on the inflammation in synoviocytes and demonstrated that leptin at its physiological concentration (1 ng/mL) promoted expressions of interleukin-6 (IL-6) and IL-8 in SW982 cells by activating p65, p38, JNK, STAT1, and STAT3. Moreover, the lowest pathological concentration of leptin potentiated the effect of IL-1β (main cytokine involved in OA pathogenesis) via activation of p65 and STAT3, leading to a significant upregulation of the IL-6 and IL-8 production. Pretreatment with omega-3 polyunsaturated fatty acids DHA and EPA suppressed the action of leptin and inhibited the IL-1β-mediated stimulation of synovitis by lowering the extent of p65 and STAT3 activation. According to our research, leptin may play a significant role in the development of OA in the joints of obese patients by promoting inflammation of synoviocytes through the activation of p65 and STAT3, while DHA and EPA, which inhibit activation of p65 and STAT3, can suppress the inflammation. Therefore, compounds that downregulate the activity of p65 and STAT3 may be the candidates for synovitis prevention and management in obese patients.

The Y-box binding protein 1 (YB-1) plays a crucial role in regulating essential cell functions, including transcription, translation, and DNA repair, through its interactions with nucleic acids and multiple protein partners. The multifunctionality of YB-1 makes the control of its levels critical for cellular homeostasis and adaptation to stress. The synthesis of YB-1 is regulated by gene transcription, protein stability (mediated by long non-coding RNAs), and translation of its mRNA. Autoregulation of YB-1 mRNA translation remains the topic of ongoing debate. Some earlier in vitro studies suggested a role of the 5′ untranslated region (UTR) in inhibiting protein synthesis, while others demonstrated the importance of YB-1 binding to the 3′ UTR for reducing translation. This disagreement has been further complicated by the absence of evidence for these mechanisms in living cells. Here, we provide the first direct evidence that YB-1 represses its synthesis in cultured human cells. Using metabolic protein labeling and immunoprecipitation, we confirmed the effect of YB-1 on the translation of its mRNA. Experiments with reporter constructs showed that both UTRs of the YB-1 mRNA are involved in autoregulation, thus resolving the contradiction in the literature. These results highlight a sophisticated mechanism for controlling YB-1 levels, which requires both 5′ and 3′ UTRs of the YB-1 mRNA, and confirm their role in fine-tuning YB-1 synthesis.