Changing Activities of the Crustacean Epidermis during the Molting Cycle

The criteria established by Drach for subdividing the moltingcycle into stages are reviewed, and a suggestion is made forimproving the uniformity of postmolt staging of different species.Changes in the epidermis during the molting cycle of the crayfishOrconectes obscurus and O. sanborni are described. EpidermalDNA content was measured throughout the cycle and found to dropsharply at stage D₀ and to rise sharply at stage A. Proteincontent declined during postmolt and rose during premolt, asexpected. Protein synthesis remained more or less constant duringpostmolt, rose during premolt, and dropped to the postmolt levelat ecdysis. Chitin synthesis appeared to follow two differentcurves depending upon whether labelled glucose or acetylglucosaminewas used as precursor. This, and the presence of a separateenzyme capable of phosphorylating acetylglucosamine and notglucose, suggests that acetylglucosamine may be utilized directlywithout prior conversion to glucose. Actinomycin D was foundto prevent increases in rate of chitin biosynthesis during premoltbut not to inhibit chitin biosynthesis already underway. Duringthe same period, actinomycin stimulated general protein biosynthesis.By utilizing the molt staging criteria described, we were ableto detect induction of premolt by ecdysone.     

Regulation of the Crustacean Mandibular Organ

The crustacean mandibular organ (MO) produces methyl farnesoate(MF), a juvenile hormone-related compound thought to have rolesin crustacean reproduction and development. Therefore, the controlof MF production by the MO has been of considerable interest.Current evidence indicates that the MO is negatively regulatedby peptides present in the eyestalk (MO inhibiting factor, MO-IH).Several eyestalk neuropeptides have been identified that inhibitMF synthesis by MO incubated in vitro. The amino acid sequencesof these MO-IH peptides are similar to peptides in the crustaceanhyperglycemic hormone (CHH) family of neuropeptides. In addition,there appears to be a compound in the eyestalk that lowers hemolymphlevels of MF in vivo but does not directly affect the MO invitro. The inhibition of MF synthesis by eyestalk peptides involvesthe inhibition of farnesoic acid O-methyl transferase, the lastenzyme in the MF biosynthetic pathway. The activity of thisenzyme is affected by cyclic nucleotides, suggesting that thesecompounds may be involved in the signal transduction pathwaymediating the effects of MO-IH.     

Ion Regulation in Crayfish: Freshwater Adaptations and the Problem of Molting

SYNOPSIS. Crayfish have a long evolutionary history in temperatefresh water (FW). Ion regulation is challenged by low externalconcentrations of Na, Cl, and Ca (<1 mM). In intermolt theprimary concern is Na and Cl balance; around ecdysis the emphasisswitches to Ca regulation as the cuticle is decalcified/calcified.Compared with marine crustaceans, intermolt crayfish maintaina reduced extracellular (EC) osmolality and have lower permeabilityto both ions and water. Hyperregulation involves active branchialuptake of Na and Cl and the unique ability to produce a hypotonicurine. Ion uptake involves apical electroneutral ion exchange(Na^$ for H^$; Cl^– for HCO₃–; counterions providedfrom CO₂ via carbonic anhydrase) followed by active basolateraltransport of Na via the Na pump, with Cl following passively.Reabsorption of 95% of filtered electrolytes at the antennalgland (kidney) involves similar subcellular mechanisms in amorphologically differentiated region of the distal tubule.Intermolt crayfish exhibit negative Ca balance (passive effluxunopposed by uptake) tolerable in view of the large cuticularCaCO₃ reserve. In premolt, cuticular Ca is reabsorbed. A smallamount is stored as gastroliths, the remainder is lost via branchialexcretion and in the discarded exuviae. At ecdysis, FW uptakegenerates the physical force for shedding, leaving the crayfishwith dilute hemolymph and a Ca deficiency. Levels of EC Na andCl are restored by intensive postmolt branchial uptake. Mineralizationof the soft exoskeleton involves remobilization of stored Caand branchial uptake of Ca and HCO₃. Transepithelial Ca transportinvolves Ca^2$ ATPase and Ca^2$/Na^$ exchange. The importance ofexternal electrolytes and pH in postmolt ion regulation is explored,as are some allometric considerations.     

Regulation of Release and Mode of Action of Crustacean Chromatophorotropins

Pigment movements in crustacean chromatophores are regulatedby pigment-concentrating and pigment-dispersing neurosecretoryhormones. The release of these hormones from the neurosecretorysystem is thought to be controlled by neural signals. Althoughthe exact cellular sources of chromatophorotropins and the natureof synaptic input to the neurosecretory system remain unresolved,recent pharmacological studies on fiddler crabs indicate thatcertain biogenic amines contribute to the release of some ofthe chromatophorotropins. The current evidence suggests that:5-hydroxytryptamine stimulates the release of RPDH (red pigmentdispersing hormone), dopamine triggers the release of RPCH (redpigment concentrating hormone), and norepinephrine stimulatesthe release of BPDH/MDH (black pigment/melanin dispersing hormone).Nothing is known of the regulation of release of leucophorotropinsand xanthophorotropins. Like most other peptide hormones, crustacean chromatophorotropinsare thought to exert their effect by first binding to receptorson the cell surface and then triggering a series of events leadingto cellular responses, but the details are far from clear. Recentwork suggests that hormone-induced pigment movements may bemodulated/mediated by alterations in cyclic nucleotide levelsand/or by changes in Ca²⁺ distribution or flux. Both cyclicnucleotides and Ca²⁺ may modulate motility by regulating theintegrity and/or function of cytoskeletal elements, but thedetails of this mechanism and the precise roles of cytoskeletalelements in the pigment-translocating process remain unresolved.     

Getting out of a Tight Squeeze--Enzymatic Regulation of Claw Muscle Atrophy in Molting

The claw closer muscle of the land crab, Gecarcinus lateralis,undergoes a cyclical atrophy and restoration during the intervalbetween ecdyses. During proecdysis (stage D₀), 30–60%of the muscle protein is degraded, which reduces tissue massand facilitates withdrawal of the propodus at ecdysis. Proteinis resynthesized as the muscle grows back to its previous sizeduring metecdysis. This atrophy is specific to the claws andcan be accentuated by multiple limb autotomy. Crustacean musclescontain five cytosolic proteinases that degrade myofibrillarproteins. Four of these constitute a family of enzymes requiringCa²⁺ for activity. These calcium-dependent proteinases (CDPs)hydrolyze myofibrillar proteins in vitro and in situ and showincreased activity in atrophic claw muscles, which suggeststhat CDPs play an important role in myofibrillar protein metabolism.The fifth enzyme is a multicatalytic proteinase (MCP), a multisubunitproteolytic complex that degrades a wide range of peptide andprotein substrates. The catalytic properties of the complexare altered with low concentrations of sodium dodecyl sulfateor by brief heating at 60°C. Only the heat-activated formdegrades myofibrillar proteins. Since the CDPs hydrolyze contractileproteins about 30-fold more rapidly than the heat-activatedMCP, the MCP probably has a more limited or specialized functionin molt-induced claw muscle atrophy.     

Functional Analysis of Insect Molting Fluid Proteins on the Protection and Regulation of Ecdysis

Molting fluid accumulates between the old and new cuticles during periodical ecdysis in Ecdysozoa. Natural defects in insect ecdysis are frequently associated with melanization (an immunity response) occurring primarily in molting fluids, suggesting that molting fluid may impact immunity as well as affect ecdysis. To address this hypothesis, proteomic analysis of molting fluids from Bombyx mori during three different types of ecdysis was performed. Many proteins were newly identified, including immunity-related proteins, in each molting fluid. Molting fluids inhibited the growth of bacteria in vitro. The entomopathogenic fungi Beauveria bassiana, which can escape immune responses in feeding larvae, is quickly recognized by larvae during ecdysis, followed by melanization in molting fluid and old cuticle. Fungal conidia germination was delayed, and no hyphae were detected in the hemocoels of pharate instar insects. Molting fluids protect the delicate pharate instar insects with extremely thin cuticles against microorganisms. To explore the function of molting fluids in ecdysis regulation, based on protein similarity, 32 genes were selected for analysis in ecdysis regulation through RNAi in Tribolium castaneum, a model commonly used to study integument development because RNAi is difficult to achieve in B. mori. We identified 24 molting proteins that affected ecdysis after knockdown, with different physiological functions, including old cuticle protein recycling, molting fluid pressure balance, detoxification, and signal detection and transfer of molting fluids. We report that insects secrete molting fluid for protection and regulation of ecdysis, which indicates a way to develop new pesticides through interrupting insect ecdysis in the future.     

Crustacean metamorphosis: an omics perspective

We quantified the independent impacts of flooding salinity, flooding depth, and flooding frequency on the native species, Phragmites australis and Scirpus mariqueter, and on the invasive species Spartina alterniflora in the Yangtze River Estuary, China. Total biomass of all three species decreased significantly with increasing salinity, but S. alterniflora was less severely affected than P. australis and S. mariqueter. Elevated flooding depth significantly decreased their live aboveground biomass of P. australis and S. mariqueter, while S. alterniflora still had high live aboveground biomass and total biomass even at the highest flooding depth. These findings indicated that S. alterniflora was more tolerant to experimental conditions than the two native species, and an unavoidable suggestion is the expansion of this non-native species in relation to the native counterparts in future scenarios of increased sea-level and saltwater intrusion. Even so, environmental stresses might lead to significant decreases in total biomass and live aboveground biomass of all three species, which would potentially weaken their ability to trap sediments and accumulate organic matter. However, the relatively high belowground-to-aboveground biomass ratio indicated phenotypic plasticity in response to stressful environmental conditions, which suggest that marsh species can adapt to sea-level rise and maintain marsh elevation.     

Molting of Rotylenchus buxophilus Golden

Molting of Rotylenchus buxophilus is similar in all observed post-embryonic stages. During molting the old cuticle and cephalic framework, conus of the old stylet, vestibulum extension, linings of amphids, distal part of the linings of the excretory duct, rectum, and phasmids are shed. The new styler is formed starting from the conus, which is simultaneously formed in its entirety.     

Preface: Recent advances in Crustacean Genomics

Aquatic plants fulfil a wide range of ecological roles, and make a substantial contribution to the structure, function and service provision of aquatic ecosystems. Given their well-documented importance in aquatic ecosystems, research into aquatic plants continues to blossom. The 14th International Symposium on Aquatic Plants, held in Edinburgh in September 2015, brought together 120 delegates from 28 countries and six continents. This special issue of Hydrobiologia includes a select number of papers on aspects of aquatic plants, covering a wide range of species, systems and issues. In this paper, we present an overview of current trends and future directions in aquatic plant research in the early twenty first century. Our understanding of aquatic plant biology, the range of scientific issues being addressed and the range of techniques available to researchers have all arguably never been greater; however, substantial challenges exist to the conservation and management of both aquatic plants and the ecosystems in which they are found. The range of countries and continents represented by conference delegates and authors of papers in the special issue illustrates the global relevance of aquatic plant research in the early twenty first century but also the many challenges that this burgeoning scientific discipline must address.     

Molting fluid chitinase: a homotropic allosteric enzyme.

The molting fluid of the tobacco hornworm has chitinase activity which shows allosteric behavior with chitin. A parabolic curve is obtained on a double reciprocal plot of $\text{1}\text{v}$ vs. $\text{1}\text{[S]}$, and a sigmoid curve results when v (N-acetylglucosamine produced) is plotted against [S] (chitin concentration in terms of N-acetylglucosamine concentration). The Hill coefficient with insect chitin is 1.95 ± 0.08. Allostery of the chitinase enables the insect to exert additional control over the integrity of structure of its cuticle.     

Physiological Control of Molting in Insects

SYNOPSIS. The initiation of a molting cycle in insects is neithera random nor a strictly periodic event. Insofar as molting canaccomplish different things under different circumstances, suchas a change in size or a change in form, it is reasonable toasume that the timing of a molt must be adapted to these functions.The onset of a metamorphic molt, in particular, must be preciselycontrolled because the onset of metamorphosis terminates thegrowth phase of a larva and establishes the body size of theadult insect. This aspect of the control of molting has receivedrelatively little attention and our knowledge of specific physiologicalmechanisms for the control of molt initiation is restrictedto three species: the blood-sucking bug, Rhodnius prolixus,the greater milkweed bug, Oncopeltus fasciatus, and the tobaccohornworm, Manduca sexta. The present review discusses the stateof our knowledge about the factors that render these insectscompetent to molt and about the stimuli that serve as a directtrigger for molting.     

Computerized Analysis of Crustacean Relationships

The strengths and weaknesses of phylogenetic analysis using computers are reviewed from the viewpoint of understanding crustacean evolution. Computerized methods require the explicit presentation of characters and character state homologies. New techniques allow investigators to design evolutionary models into a character data matrix, or to use evolutionary models that make minimal a priori assumptions. The computer analysis relieves the investigator from the highly repetitious testing of trees, allows the concentration on the character state data, and provides objective methods for comparing trees, primarily their length. These are regarded as the strengths of computerized methods. The weaknesses of these methods include the relatively inscrutable nature of the character data matrix compared with the overall ‘gestalt’ of resulting trees, the difficulties of defining discrete homologies within the Crustacea, especially for counts of segmentation, the lack of clear intermediate character states in some multistate segmental characters, and the inability to define evolutionary polarity. These difficulties may be overcome by analysing the data using the minimal assumption models of character evolution, and by a recognition that the trees are a result of the input data, and therefore the data should be criticized, rather than the trees themselves. A ‘consensus’ character data set, including most extant major groups of the Crustacea as well as several key fossils, was assembled and revised by the participants in the workshop. An artificial taxon, ‘ur-crustacean characters’, was introduced to root the tree. Three observations may be made from parsimony analyses using several weighting and tree rooting methods. (1) The currently accepted large scale phylogeny and classification of the Crustacea is not corroborated. (2) The number of supposed plesiomorphic traits possessed by a taxon is not a good index for early derivation in crustacean evolution. (3) The taxon Maxillopoda is not supported by the arrangement of any of the trees.     

Crustacean Vitellogenesis: Its Role in Oocyte Development

One of the major changes that occurs during the maturation ofoocytes is the accumulation of yolk protein, or vitellin (Vn).To better understand how this process is regulated, we characterizedthe Vn of the ridgeback shrimp, Sicyonia ingentis (Penaeoidea).This Vn is a 322 kDa molecule composed of three subunits. Usingpurified Vn, we developed an anti-Vn antiserum and used it tocharacterize vitellogenin by Western blot analysis. The antiserumwas also used in an ELISA to measure hemolymph levels of vitellogenin.Previous studies suggested the presence of vertebrate-type steroidsmight stimulate reproductive processes in decapod crustaceans.Treatment of sexually quiescent female shrimp with progesterone,hydroxyprogesterone, and estradiol did not increase hemolymphlevels of yolk protein precursor. The absence of a responseto these steroids may reflect the presence of other hormones(such as the gonad-inhibiting hormone) that prevent oocyte development.To examine the molecular basis for the regulation of vitellogenesis,ovarian and hepatopancreas expression cDNA libraries were screenedusing the anti-Vn antiserum. A 2.9 kilobase clone was isolatedfrom both cDNA libraries suggesting that both tissues are sitesof vitellogenin synthesis. These molecular tools should be usefulfor in vitro studies of vitellogenin synthesis.     

Phocanema decipiens: Molting in seals

Infective larvae of the anisakine nematode Phocanema decipiens from the muscle of Atlantic cod (Gadus morhua) were fed to harbor seals (Phoca vitulina) and gray seals (Halichoerus grypus). During maturation in the stomach of seal hosts, P. decipiens molted twice; these molts are the third and fourth of its life cycle. The third molt occurred between the second and fifth days of infection. The third stage, i.e., infective larva entering the third molt, had a cuticular tooth ventral to the mouth; the fourth stage larva emerging from the third molt had three bilobed lips with dentigerous ridges. The fourth molt occurred between the 5th and 15th days in seals. A female nematode emerging from the fourth molt possesses a vulva and a vagina; a male possesses caudal alae, pre- and postanal papillae. Significant morphometric changes in nematodes were associated with both molts. Females and males of P. decipiens reached maturity after 15 to 25 days in seals. Ova were detected in the feces of the seal hosts as early as the 16th day.     

Cyclin-CDK-CKI及UPP参与生殖调控及在甲壳动物性腺发育中的研究进展

Cyclin-CDK-CKI是参与真核细胞周期调控的3个重要的调节因子。泛素蛋白酶体途径(ubiquitin proteasome pathway,UPP)是真核细胞内非溶酶体途径的蛋白质选择性降解的重要途径。细胞周期中许多时相特异性周期蛋白通过泛素化被周期性的降解对于细胞分裂的调控进程具有重要的作用。主要综述了Cyclin-CDK-CKI和UPP的组成、生理生化特性及其参与生殖调控的作用机制,同时阐述了这两大系统之间的联系及其在甲壳动物性腺发育中的研究进展。     

Structure and Function of the Transverse Tubular System in Crustacean Muscle Fibers

The structure and function of the transverse tubular system(TTS) in two types of crustacean muscle fibers are examined.Giant fibers from the barnacle,Balanus nubilus, which are gradedlycontracting, are compared with allor-none twitch fibers fromthe crab, Carcinus maenas. Both fiber types were found to havedeep sarcolemmal invaginations which serve both to increasethe fiber surface area and to kfeep the length of the tubulesshort enough for electrotonic propagation.The ultrastructureof the tubular system in both types of fiber is compared.Thesystem is better developed in Carcinus than in Balanus, butthe slow Balanus fibers do have a relatively well developedTTS and sarcoplasmic reticulum in contrast to slow vertebratefibers. The apparent high, membrane-capacitance values of crustaceanfibers are the result of investigators not taking into considerationthe large increase in surface area due to the sarcolemmal infoldings.Thetubular membranes in Carcinus fibers were found to be permselectiveto chloride ions, and could be made to swell (as confirmed byelectron microscopy) by establishing an outward gradient forchloride across them. The capacitance of the tubular membranerelative to the plasma membrane was found to increase when thetubuleswere swollen. The implication of a fiber having two spatiallyseparated, differentially permeable membranes on excitation-contractioncoupling is discussed.