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At least 19 recordsLinked to original sources
PubMed2026

Cryopreservation-induced proteomic alterations in Pêga donkey (Equus asinus) spermatozoa.

Semen cryopreservation is a key tool for assisted reproduction and genetic conservation, but its efficiency remains limited in donkeys, compromising post-thaw sperm quality. This study aimed to characterize the proteomic profile of Pêga donkey spermatozoa and to investigate molecular mechanisms associated with cryopreservation-induced impairment of sperm function. Semen samples were collected from Pêga jacks and evaluated for sperm motility and vigor before and after cryopreservation. Quantitative proteomic analysis was performed by LC-MS/MS, followed by bioinformatic characterization of differentially abundant proteins. Cryopreservation markedly reduced sperm motility in all animals, whereas sperm vigor showed only a non-significant tendency toward reduction, suggesting sublethal cryoinjury primarily affecting flagellar efficiency. Proteomic profiling identified 554 proteins, of which 98 were differentially abundant between in natura and cryopreserved spermatozoa. Functional enrichment analyses showed that these proteins were mainly associated with energy metabolism, mitochondrial oxidative phosphorylation, glycolysis, cytoskeletal organization, signal transduction, proteostasis, and oxidative stress response. Notably, proteins involved in ATP production, mitochondrial function, and axonemal organization were significantly altered, supporting a mechanistic link between metabolic dysfunction, flagellar structural disorganization, and reduced post-thaw motility. Overall, cryopreservation induced coordinated and compartment-specific remodeling of the donkey sperm proteome, particularly affecting pathways essential for motility and functional competence. These findings provide new molecular insights into the cryobiological vulnerability of donkey spermatozoa and establish a mechanistic basis for the development of more effective, biology-driven cryopreservation strategies for this species.

Animals↗
PubMed2026

Biallelic Variants in ATP1A4 Are Associated with Oligoasthenoteratozoospermia and Male Infertility.

Male infertility, often caused by structural and functional sperm defects, remains genetically unexplained in a substantial proportion of cases. ATP1A4 encodes a testis-specific isoform of the Na+, K+-ATPase, a membrane enzyme crucial for maintaining cellular ionic homeostasis. Previous studies on Atp1a4 knockout mice have demonstrated severe defects in sperm motility and flagellar architecture; however, the contribution of ATP1A4 variants to human male reproduction remains to be elucidated. In this study, we identified compound biallelic variants in ATP1A4, a missense variant (c.2578 T>A, p.Tyr860Asn) and a frameshift variant (c.2582del, p.Gly861Aspfs*5), in a patient presenting with severe oligoasthenoteratozoospermia. Both variants markedly affected ATP1A4 protein expression. Morphological analyses revealed coiled and folded flagella, disrupted mitochondrial sheaths, and irregular head morphology in the patient's spermatozoa. Expression profiling revealed that ATP1A4 was highly enriched in post-meiotic spermatids and localized along the entire flagellum of mature sperm in both humans and mice, indicating a critical role in flagellar assembly and structural integrity. Notably, intracytoplasmic sperm injection (ICSI) in this patient resulted in low fertilization efficiency and failed implantation, suggesting a potential adverse impact of ATP1A4 deficiency on sperm functional competence beyond motility. These findings broaden the genetic spectrum of oligoasthenoteratozoospermia and highlight ATP1A4 as a potential gene associated with human male infertility.

Male↗
PubMed1979

Structure, protein composition and birefringence of the costa: a motile flagellar root fibre in the flagellate Trichomonas.

The costa is a rod-shaped intracellular organelle lying in the cytoplasm immediately below the undulating membrane. In certain large species of Trichomonas (T. gigantea, T. termopsidis and an undescribed species from the termite Porotermes adamsoni) this organelle is motile. Bending waves are transmitted along the length of the costa, in either direction. It is shown that the bending is actively produced by the costa itself. The form of the movements is described in detail. The costa is birefringent. At the point at which bending occurs there is a sharply localized reduction in birefringence. Electron microscopy shows that the costa is composed of longitudinally running lamellae, 2--3 nm thick and spaced 12 nm apart. These are connected to transversely running elements spaced at intervals of about 37 nm. The lamellae occur in two alternative configurations: straight and zig-zag. Bending probably results from a local, transient change from the straight to the zig-zag configuration. This would account for the local change in birefringence which accompanies bending. Polyacrylamide gel electrophoresis of isolated costas shows them to contain a number of protein components, of which the principal one has a molecular weight of about 90,000. Preliminary cytochemical evidence is presented for ATPase activity in the costa. The costa is a type of motile system distinct from any hitherto described.

Adenosine Triphosphatases↗
PubMed1976

Antigenic structures of Salmonella flagella. I. Presence of an antigenic determinant exposed at one end of flagellar fragments.

Salmonella flagellin, which is a constitutional subunit of the flagellum, was shown to have antigenic determinants distinct from its own serotypic ones. These antigenic determinants were found to be common to flagellins from the so-called g-complex serotypes, such as fg, mt, gm, gt, gp and gmptu, but not to those from other serotypes, such as a, i or enx. Rabbits immunized with flagellin of serotype "fg" produced anti-"fg" flagellin antibodies. Only about 20 percent of these corresponded to the serotype determinants of the "fg" on the surface of the flagella, and the remaining 80 percent reacted with the flagellin of the unrelated serotype "mt", and corresponded to the distinct determinants common to the flagellin molecules. These antigenic determinants were detected by the immunoferritin technique at only one, not both, terminals of the flagellar fragments, suggesting that a unidirectional arrangement of flagellin subunits in the flagella may expose the inherent conformation of the subunits at only one end of the flagellum.

Antibodies, Bacterial↗
PubMed2025

Structural properties of short-chain carboxylic acids and alcohols relate to the molecular and physiological response of Salmonella enterica in an acidic environment.

Short-chain carboxylic acids (SCCA) and short-chain alcohols (SCALC) are naturally occurring antimicrobials that contribute to the biopreservation of food fermentations. This study investigated the effect of structurally different SCCA/SCALC with two-carbon (acetic acid; phenylacetic acid; 2-phenylethanol), three-carbon (propionic acid; 3-phenylpropionic acid; 3-phenylpropanol), and three-carbon chain with an additional hydroxyl group (lactic acid; 3-phenyllactic acid; 1-phenylpropanol) on the fitness, metabolic activity and gene expression of the pathogen Salmonella enterica at pH 4.5. SCCA inhibited Salmonella at lower concentrations than SCALC with the exception of lactic acid, which was partly consumed. The presence of a phenyl group enhanced antimicrobial activity. SCCA but not SCALC increased the lag phase of S. enterica, and in general, acetate was formed when cell growth was reduced by 20% suggesting a negative impact on bacteria fitness. Principal component analysis and hierarchical clustering indicated distinct gene expression profiles of S. enterica in response to SCCA or SCALC. In the presence of certain SCCA/SCALC, Salmonella activated pathways related to cellular pH control, and 1,2-propanediol, propionic acid and ethanolamine metabolism that involved the formation of metabolosomes. Genes related to flagellar assembly were less expressed and mobility was lower in the presence of lactic and 3-phenyllactic acid compared to controls suggesting a compound-specific response. KEY POINTS: • Differences in response among structurally different SCCA/SCALC at acidic condition. • SCCA/SCALC stress interfered with cell growth and metabolism of acetic and propionic acid. • Lactic acid prolonged the lag phase and reduced motility of Salmonella.

Salmonella enterica↗
PubMed1979

Cytochemical analysis at the fine-structural level of trypanosomatids stained with phosphotungstic acid.

The ethanolic phosphotungstic acid (PTA) technic was used to detect, at the fine-structural level, basic proteins in various developmental stages of pathogenic Trypanosoma cruzi, and nonpathogenic Herpetomonas samuelpessoai, Leptomonas samueli, and Crithidia deanei, trypanosomatids. Reactions were observed in the nucleus of all stages. In the kinetoplast of epimastigote and promastigote forms reactions were noted mainly at the periphery. In trypomastigotes and choanomastigotes forms, however, an intense reacion was observed thorughout the kinetoplast. Reactions were present in cytoplasmic vesicles related to protein storage in T. cruzi and in membrane-bounded peroxisome-like organelles of H. samuelpessoai, L. samueli and C. deanei. The network of filaments which forms the paraxial rod did not react. In the flagellum, reaction was noted only at the peripheral doublet microtubules. PTA reacts also with structures related to the junction between the flagellar and cell body membranes.

Animals↗
PubMed1979

Regulation of expression of the flagellin gene (hag) in Escherichia coli K-12: analysis of hag-lac gene fusions.

Previous studies have defined 28 genes necessary for the synthesis of the flagellar apparatus of Escherichia coli K-12. This study analyzed the influence of the flagellar genes on the expression of the hag gene (structural gene for flagellin). To this end, a hag::Mu d(Apr lac) mutant which had the lac genes fused to the promoter of the hag gene was constructed. This allowed the measurement of hag gene expression by detection of beta-galactosidase activity. The following observations were made. (i) The hag gene was expressed constitutively in Fla+ cells. (ii) hag gene expression was positively regulated by flaA, FLAB, flaC, flaD, flaE, flaG, flaH, flaI, flaK, flaL, flaM, flaN, flaO, flaP, flaQ, flaR, flaV, flaW, flaX, flaY, flaZ, flbA, and flbB genes.hag-lac expression was not observed in strains with these fla mutations. (iii) The hag gene was expressed in mutants with flaS, flaT, flaU, and flbC defects. Therefore, these genes were not involved in regulation of hag gene transcription.

Bacterial Proteins↗
PubMed1978

Isolated flagellar apparatus of Chlamydomonas: characterization of forward swimming and alteration of waveform and reversal of motion by calcium ions in vitro.

The control of flagellar activity in the biflagellate green alga, Chlamydomonas reinhardtii was investigated by the in vitro reactivation of the isolated flagellar apparatus (the 2 flagella attached to their respective basal bodies plus accessory structures). The waveform and beat frequency of the isolated apparatus in the presence of 1 mM adenosine triphophate (ATP) were comparable to those recorded for living cells. Equimolar concentrations of adenosine diphosphate (ADP) could be substituted for ATP with little change in beat frequency and no apparent change in waveform, suggesting that the latter is converted to ATP by axonemal adenylate kinase. No reactivation occurred in adenosine monophosphate (AMP). But frequencies in cytidine, guanosine and uridine triphosphates (CTP, GTP and UTP) were approximately 10% that obtained in ATP. Reactivation was optimal over a broad pH range between pH 6.4 and pH 8.9 in both APT and ADP. Isolated flagellar apparatus could be induced to change from forward to reverse motion in vitro by manipulation of exogenous calcium ions. The 2 types of motion were directly comparable to recorded responses of living cells. Forward swimming occurred at levels of calcium below 10(-6)M, the isolated apparatus changing to backward motion above this level. Motility was inhibited at concentrations above 10(-3)M. The threshold for reversal of motion by calcium was lowered to 10(-7)M when the flagellar membranes were solubilized with detergent, indicating that the flagellar membranes are involved in the regulaion of the level of calcium within the axoneme. The reversal of motion by calcium was itself freely reversible. The relationship of these observations to the known tactic responses of Chlamydomonas is discussed.

Adenosine Triphosphate↗
PubMed2025

Laboratory Evolution Reveals Transcriptional Mechanisms Underlying Thermal Adaptation of Escherichia coli.

Adaptive laboratory evolution is able to generate microbial strains, which exhibit extreme phenotypes, revealing fundamental biological adaptation mechanisms. Here, we use adaptive laboratory evolution to evolve Escherichia coli strains that grow at temperatures as high as 45.3 °C, a temperature lethal to wild-type cells. The strains adopted a hypermutator phenotype and employed multiple systems-level adaptations that made global analysis of the DNA mutations difficult. Given the challenge at the genomic level, we were motivated to uncover high-temperature tolerance adaptation mechanisms at the transcriptomic level. We employed independently modulated gene set (iModulon) analysis to reveal five transcriptional mechanisms underlying growth at high temperatures. These mechanisms were connected to acquired mutations, changes in transcriptome composition, sensory inputs, phenotypes, and protein structures. They are as follows: (i) downregulation of general stress responses while upregulating the specific heat stress responses, (ii) upregulation of flagellar basal bodies without upregulating motility and upregulation fimbriae, (iii) shift toward anaerobic metabolism, (iv) shift in regulation of iron uptake away from siderophore production, and (v) upregulation of yjfIJKL, a novel heat tolerance operon whose structures we predicted with AlphaFold. iModulons associated with these five mechanisms explain nearly half of all variance in the gene expression in the adapted strains. These thermotolerance strategies reveal that optimal coordination of known stress responses and metabolism can be achieved with a small number of regulatory mutations and may suggest a new role for large protein export systems. Adaptive laboratory evolution with transcriptomic characterization is a productive approach for elucidating and interpreting adaptation to otherwise lethal stresses.

Escherichia coli↗
PubMedDate not supplied

[Electron microscopic study of presumptive photoreceptor cells in the aboral organ of the ctenophore, Beroë cucumis].

Presumptive photoreceptor cells are located in the receptor epithelium of the aboral organ and exhibit some common morphological characters with flagellar photoreceptors of some invertebrates, as well as with photoreceptors of vertebrates. These cells are specifically distinguished by the presence of a special membrane structure in their epinuclear zone, this structure being the derivative of a cilium. Presumptive photoreceptor cells which have a short central projection and form in their basal part the synaptic contacts ("triads") with the adjacent nervous endings, may be classified as pseudosecondary receptor cells. Acetylcholinesterase was found in the synaptic regions. Cytoplasm of the presumptive photoreceptor cells is rich in granular vesicles which contain biogenic amines.

Animals↗
PubMed1976

Filaments of Trypanosoma brucei: some notes on differences in origin and structure in two strains of Trypanosoma (Trypanozoon) brucei rhodesiense.

Filaments attached to trypanosomes of two strains of T. (T.) brucei were studied by electron microscopy and two distinct types identified: short-thick and long-thin. The former are associated with stumpy trypanosomes and are secretions, via the flagellar pocket, which originate in the area of the Golgi complex, during the infection of the host. They are referred to as 'secretory filaments'. Their diameter is 0.09 to 0.14 mum. The long-thin filaments are associated with slender forms of trypanosome in various artificial situations; those shown by negative staining are believed to be cytoplasmic extrusions from the anatomically weak extremities of the parasite and are referred to as 'plasmanemes'. Their diameter is 0.06 mum. Both types appear to maintain their structure without the aid of the normal type of unit membrane as myelin formations.

Animals↗
PubMed1975

[A functional flagella with a 6 + 0 pattern].

The male gamete of the Gregarine Lecudina tuzetae has been studied with transmission electron microscopy and microcinematography. It is characterized by a flagellar axoneme of 6 + 0 pattern, a reduction of the chondriome, and the abundance of storage polysaccharide or lipid bodies. The movements of the flagella are of the undulating type and they are performed in the three dimensions of space. They are very slow, with a cycle time of about 2s. The structure of the axoneme components are similar to those of flagella with a 9 + 2 pattern. Each doublet has overall dimensions of 350 x 220 A; the space between the adjacent doublets is about 160 A. The A subfiber bears arms like dynein arms. The diameter of the axoneme is about 1,000 A. The basal body consists of a cylinder of dense material 2,500 A long and 1,300-1,400 A in diameter; a microtubule 200 A in diameter is present in the axis. This study shows that a 6 + 0 pattern can generate a flagellar movement. The mechanism of the flagellar movement of the male gamete of L. tuzetae does not require the presence of central microtubules and it would include molecular interactions of the dynein-tubulin type between the adjacent peripheric doublets. The slowness of the movements is discussed in terms of the axoneme's structure and its energy supply. Finally, the phylogenetic significance of this flagella is examined on the basis of the morphopoietic potentialities of the centriolar structures.

Animals↗
PubMed2004

Ultrastructure of Proteus mirabilis swarmer cell rafts and role of swarming in catheter-associated urinary tract infection.

Proteus mirabilis is a common cause of catheter-associated urinary tract infection (C-UTI). It blocks indwelling urethral catheters through the formation of extensive crystalline biofilms. The obstruction of urine flow can induce episodes of pyelonephritis, septicemia, and shock. P. mirabilis exhibits a type of motility referred to as swarming, in which multicellular rafts of elongated, hyperflagellated swarmer cells form and move rapidly in concert over solid surfaces. It has been suggested that swarming is important in the pathogenesis of C-UTI. In this study we generated a set of stable transposon mutants deficient in swarming and used them to assess the role of swarming in the migration of P. mirabilis over urinary catheters. Swarming was found to be essential for migration over all-silicone catheters. Swarming-deficient mutants were attenuated in migration over hydrogel-coated latex catheters, but those capable of swimming motility were able to move over and infect these surfaces. A novel vapor fixation technique for the preparation of specimens and scanning electron microscopy were used to resolve the ultrastructure of P. mirabilis multicellular rafts. The flagellar filaments of P. mirabilis were found to be highly organized during raft migration and were interwoven in phase to form helical connections between adjacent swarmer cells. Mutants lacking these novel organized structures failed to swarm successfully. We suggest that these structures are important for migration and formation of multicellular rafts. In addition, the highly organized structure of multicellular rafts enables P. mirabilis to initiate C-UTI by migration over catheter surfaces from the urethral meatus into the bladder.

DNA Transposable Elements↗
PubMed2026

Motile and non-motile Listeria species adopt distinct ecological and evolutionary strategies to achieve broad geographic ranges across soil ecosystems.

Broad geographic ranges often reflect ecological versatility and are associated with lower extinction risk. Motility is a key physiological and ecological trait in bacteria. However, how some motile and non-motile bacteria achieve broad geographic ranges remains poorly understood. Here, we analyzed the genomes of 141 Listeria welshimeri and 90 Listeria booriae isolates systematically obtained from soils, representing widespread motile and non-motile species, respectively. We show that L. welshimeri lacks clear phylogeographic structure, suggesting minimal geographic barriers to dispersal. Its wide distribution is likely associated with enhanced motility and effective host colonization that facilitate wildlife-driven dispersal, particularly by regional-terrestrial birds. This pattern is supported by positive selection on flagellar and chemotaxis genes, strong associations with wildlife movement patterns, and close genomic relatedness between soil and wild bird isolates. In contrast, L. booriae displays clade endemism and a strong distance-decay relationship, suggesting dispersal limitation. Despite lacking a dispersal advantage, L. booriae's wide distribution appears to be linked to genomic flexibility and metabolic versatility that support adaptation to diverse environmental conditions, especially those shaped by iron concentration and precipitation. This is evidenced by its large, open pangenome characterized by abundant and diverse metabolic pathways and broad substrates utilization capacity; pronounced positive selection on genes involved in inorganic ion, amino acid, and coenzyme transport and metabolism; and strong associations between gene richness and abiotic factors as well as bacterial community composition. These findings suggest distinct genomic foundations and ecological and evolutionary mechanisms underlying the success of motile and non-motile cosmopolitan bacteria in soil ecosystems.

Soil Microbiology↗
PubMed1977

Identification of dynein as the outer arms of sea urchin sperm axonemes.

The location of dynein, the main flagellar ATPase, within the sea urchin sperm axoneme was investigated by the use of immunofluorescence and immunoelectron microscopy, employing an antiserum against a tryptic fragment of dynein 1 (Fragment 1A) purified from sea urchin sperm flagella. The axonemes were found to be stained with the antiserum when examined by an indirect immunofluorescence technique. Immunoelectron microscopy with the antiserum and a ferritin-conjugated IgG fraction of goat antiserum to rabbit IgG revealed that, among the structures within the axoneme, only the outer arms were labeled with ferritin particles. With either the normal serum or antiserum absorbed with Fragment 1A, there were no ferritin particles within the axonemes. When the outer arms were extracted with 0.5 M NaCl, leaving the inner arms intact, again no ferritin dots were detected. Furthermore, it was found that the outer arm on the no. 5 doublet microtubule, which connects with the extra arm projection backward from the no. 6 doublet, had no attached ferritin particles. From these observations, it can be concluded that the outer arm consists of dynein (at least dynein 1) and that Fragment 1A, containing the active site for ATPase activity of dynein 1, is located at the distal end of the outer arms. The significance of the present findings is considered in connection with flagellar movement.

Adenosine Triphosphatases↗
PubMed1979

Study of the mechanism of vanadate inhibition of the dynein cross-bridge cycle in sea urchin sperm flagella.

The effect of vanadate on the ATP-induced disruption of trypsin-treated axonemes and the ATP-induced straightening of rigor wave preparations of sea urchin sperm was investigated. Addition of ATP to a suspension of trypsin-treated axonemes results in a rapid decrease in turbidity (optical density measured at 350 nm) concomitant with the disruption of the axonemes by sliding between microtubules to form tangles of connected doublet microtubules (Summers and Gibbons, 1971; Sale and Satir, 1977). For axonemes digested to approximately 93 percent of their initial turbidity, 5 {muM} vanadate completely inhibits the ATP-induced decrease in turbidity and the axonemes maintain their structural integrity. However, with axonemes digested to approximately 80 percent of their initial turbidity, vanadate fails to inhibit the ATP-induced decrease in turbidity and the ATP-induced structural disruption of axonemes, even when the vanadate concentration is raised as high as 100 mum. For such axonemes digested to 80 percent of their initial turbidity, the form of ATP-induced structural changes, in the presence of 25 muM vanadate, was observed by dark-field light microscopy and revealed that the axonemes become disrupted into curved, isolated doublet microtubules, small groups of doublet microtubules, and "banana peel" structures in which tubules have peeled back from the axoneme. Addition of 5 muM ATP to rigor wave sperm, which were prepared by abrupt removal of ATP from reactivated sperm, causes straightening of the rigor waves within 1 min, and addition of more than 10 muM ATP causes resumption of flagellar beating. Addition of 40 muM vanadate to the rigor wave sperm does not inhibit straightening of the rigor waves of 2 muM-1 mM ATP, although oscillatory beating is completely inhibited. These results suggest that vanadate inhibits the mechanochemical cycle of dyein at a step subsequent to the MgATP(2-)-induced release of the bridged dynein arms.

Adenosine Triphosphatases↗
PubMed2025

Spatio-genetically coordinated TPR domain-containing proteins modulate c-di-GMP signaling in Vibrio vulnificus.

Vibrio species, which include several pathogens, are autochthonous to estuarine and warm coastal marine environments, where biofilm formation bolsters their ecological persistence and transmission. Here, we identify a bicistronic operon, rcbAB, whose products synergistically inhibit motility and promote biofilm maturation post-attachment by modulating intracellular c-di-GMP levels in the human and animal pathogen V. vulnificus. RcbA contains an N-terminal tetratricopeptide repeat (TPR) domain and a structured C-terminal region of unknown function, while RcbB possesses an N-terminal TPR domain and a C-terminal GGDEF domain characteristic of diguanylate cyclases. The TPR domain of RcbB represses its diguanylate cyclase activity, while RcbA's TPR domain and C-terminal region co-operatively de-repress it. Localization of both proteins to the flagellar pole is TPR-dependent but not co-dependent, although RcbA anchors RcbB to the pole in the absence of polar landmarks such as HubP and flagella. The conservation of rcbAB across diverse bacterial taxa substantiates its fundamental importance in bacterial biology. This work demonstrates how spatio-genetically coordinated TPR domain-containing proteins modulate c-di-GMP signaling, contributing to our understanding of biofilm formation in Vibrio species and potentially other bacteria. It also reveals the first evidence of inter-protein interaction via the TPR domains of both partners, challenging the conventional paradigm in which only one bears the domain.

Vibrio vulnificus↗