An Embryonic Diapause-like Adaptation with Suppressed Myc Activity Enables Tumor Treatment Persistence

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Maternal GABAergic and GnRH/corazonin pathway modulates egg diapause phenotype of the silkworm Bombyx mori

Diapause represents a major developmental switch in insects and is a seasonal adaptation that evolved as a specific subtype of dormancy in most insect species to ensure survival under unfavorable environmental conditions and synchronize populations. However, the hierarchical relationship of the molecular mechanisms involved in the perception of environmental signals to integration in morphological, physiological, behavioral, and reproductive responses remains unclear. In the bivoltine strain of the silkworm Bombyx mori, embryonic diapause is induced transgenerationally as a maternal effect. Progeny diapause is determined by the environmental temperature during embryonic development of the mother. Here, we show that the hierarchical pathway consists of a γ-aminobutyric acid (GABA)ergic and corazonin signaling system modulating progeny diapause induction via diapause hormone release, which may be finely tuned by the temperature-dependent expression of plasma membrane GABA transporter. Furthermore, this signaling pathway possesses similar features to the gonadotropin-releasing hormone (GnRH) signaling system for seasonal reproductive plasticity in vertebrates.

To ensure survival under unfavorable environmental conditions and synchronize populations, most insect species enter diapause, which is a seasonal adaptation that evolved as a specific subtype of dormancy (1, 2). Diapause is not a passive response to changing conditions but rather an actively induced state that precedes adverse natural situations. Therefore, this diapause phenotype is accompanied by changes in energy metabolism or storage to improve cold/stress tolerance in later life stages, or progeny via reproductive switch (3). Although it has been generally suggested that brain/neuroendocrine systems are associated with this seasonal reproductive plasticity in both vertebrates and invertebrates (3, 4), the hierarchical relationship of the molecular mechanisms involved in the perception of environmental signals to integration into morphological, physiological, behavioral, and reproductive responses, known as the diapause syndrome, remains unclear (3).The silkworm Bombyx mori is a typical insect that arrests normal development during early embryogenesis, which is accompanied by metabolic changes in diapause (5, 6). The development of diapause-destined embryos is arrested during the G2 cell cycle stage immediately after the formation of the cephalic lobe and telson and sequential segmentation of the mesoderm (7). The bivoltine strain of B. mori has two generations per year, and progeny diapause is transgenerationally induced as a maternal effect and is determined by the environmental temperature, photoperiod, and nutrient conditions during embryonic and larval development of the mother (5, 6). The temperature signal during the mother’s embryonic development predominantly affects diapause determination, even if silkworms of the bivoltine Kosetsu strain are exposed to all cases of photoperiods during embryonic and larval development. In the Kosetsu strain, when eggs are incubated at 25 °C under continuous darkness, the resultant female moths (25DD) lay diapause eggs in almost all cases. In contrast, incubation of eggs at 15 °C in dark condition results in moths (15DD) that lay nondiapause eggs in almost all cases (6).Embryonic diapause is induced by the diapause hormone (DH) signaling pathway, which consists of highly sensitive and specific interactions between a neuropeptide, DH, and DH receptor (DHR) (6, 8). DH is exclusively synthesized in seven pairs of neurosecretory cells (DH-PBAN–producing neurosecretory cells [DHPCs]) located within the subesophageal ganglion (SG) in the mother’s generation (6). DH is released into the hemolymph during pupal–adult development and acts on the DHR, which belongs to the G protein-coupled receptors (GPCRs) (9). DH levels in the hemolymph are higher in the 25DD than 15DD pupae in the middle of pupal–adult development when the developing ovaries are sensitive to DH (6). Furthermore, the embryonic Bombyx TRPA1 ortholog (BmTRPA1) acts as a thermosensitive channel that is activated at temperatures above ∼21 °C and affects diapause induction through DH release (10). However, there remain questions about the thermal information that is received by BmTRPA1 and linked to DH signaling to induce diapause.From the 1950s, it has been suggested that the DH release was controlled by signals derived from certain region(s) in the brain based on surgical experiments, such as midsagittal bisection or transection (11–13). Especially, the operation in nondiapause producers changed them to diapause producers while transection of the protocerebrum had no effect on the diapause producers. These surgical results suggested the involvement of the protocerebrum in the inhibitory control of DH secretion (12, 14). Furthermore, the accumulation of the ovarian 3-hydroxykynurenine (3-OHK) pigment that accompanies the diapause syndrome was affected by injection with γ-aminobutyric acid (GABA) and the plant alkaloid picrotoxin (PTX), which is a widely used ionotropic GABA and glycine receptor antagonist (15, 16), and the selective ionotropic GABA receptor (GABAR) antagonist bicuculline. This suggests that a GABAergic neurotransmission via ionotropic GABAR is involved in DH secretion, which may be active in nondiapause producers but inactive in diapause producers throughout the pupal–adult development (14, 17). In general, ionotropic GABAR is composed of homo- or hetero-pentameric subunits. All known GABAR subunits display a similar structural scheme, with a large N-terminal extracellular domain involved in the formation of a ligand-binding pocket and a pore domain made of four transmembrane alpha-helices (TM1–TM4) (16, 18). Four homologous sequences of the ionotropic GABAR subunit genes were identified as RDL, LCCH3, GRD, and a GRD-like sequence named 8916 in various insects (19). However, the in vivo physiological roles of both signals derived from the brain and the GABAergic pathway in diapause induction have not been previously investigated.Corazonin (Crz) is an undecapeptide neurohormone sharing a highly conserved amino acid (a.a.) sequence across insect lineages and is involved in different physiological functions, such as heart contraction (20), stress response (21, 22), various metabolic activities (23–25), female fecundity (26), melanization of locust cuticles (27), regulation of ecdysis (28, 29), and control of caste identity (30). Moreover, Crz belongs to the gonadotropin-releasing hormone (GnRH) superfamily alongside adipokinetic hormone (AKH) and AKH/Crz-related peptide (ACP). Duplicates of an ancestral GnRH/Crz signaling system occurred in a common ancestor of protostomes and deuterostomes through coevolution of the ligand receptor (31, 32).Herein, we demonstrated that the hierarchical pathway consists of a GABAergic and Crz signaling system modulating progeny diapause induction by acting on DH release. We propose that the PTX-sensitive GABAergic signal may act to chronically suppress Crz release in dorsolateral Crz neurons (under nondiapause conditions) and that diapause conditions (or PTX injection) inhibits GABAergic signaling, resulting in accelerated Crz release, which in turn induces DH release. GABA signaling may be finely tuned by the temperature-dependent expression of the plasma membrane GABA transporter (GAT), which differs between the 25DD and 15DD conditions. Furthermore, this signaling pathway possesses similar features to the GnRH signaling system with respect to seasonal reproductive plasticity in vertebrates.     

超声诊断早期胚胎停育

目的探讨彩色多普勒超声在早期胚胎停育中的诊断价值。方法应用经腹及经阴道彩色多谱勒超声对首次超声确诊及可疑胚胎停育二次超声确诊病例的超声表现进行分析总结。结果超声一次确诊者60例（80％）,二次确诊者15例（20％）,其中枯萎孕卵33例（45％）,均未见卵黄囊显示;胚胎死亡21例（28%）,其中卵黄囊枯萎17例（23%）,增大4例（5．3％）。结论经腹及经阴道彩色多普勒超声能对胚胎停育及早、快速、准确地进行明确诊断,能早期发现宫内胚胎发育异常,可指导临床及时处理,具有重要的临床应用价值。     

Expression of the Hsp83 gene in response to diapause and thermal stress in the moth Sesamia nonagrioides

A full-length Hsp83, named SnoHsp83, cDNA from the corn stalk borer, Sesamia nonagrioides, was cloned and sequenced. Genomic analysis showed that the SnoHsp83 gene is unique. The size of the SnoHsp83 cDNA was found to be approximately 2.6 kb. The deduced polypeptide comprised 717 amino acid residues, with a molecular mass of 82.6 kDa. It contained all the highly conserved amino acid motifs that characterize the cytosolic members of the hsp90 family. We investigated the expression of SnoHsp83 gene in response to diapause and heat/cold stress. SnoHsp83 is constitutively expressed in non-diapausing larvae and is induced 15-fold by heat. SnoHsp83 displays a similar pattern to SnoHsc70 under diapause conditions, when extra larval moults occur. Our results indicate that the SnoHsp83 gene could be involved in the developmental process that occurs between two moults.     

Do human concepti have the potential to enter into diapause?

Although there is no direct evidence as to whether human concepti have the potential to enter into diapause before implantation, the possibility that human concepti may be capable of following this developmental pathway if exposed to an appropriate environment cannot be ruled out. Direct evidence remains elusive because of the ethical restraints associated with research activities within this area of knowledge. If conceptus diapause has evolved in primates and persists at the present time despite its apparent limited or no adaptive advantage, artificial induction of diapause in humans may have clinical implications for increasing: (i) the viability of concepti after biopsy, freezing-thawing or any other experimental procedure that tends to decrease cell numbers or division rate of concepti; and (ii) the relatively low implantation rates obtained at the present time after uterine transfer of human concepti fertilized in vitro. Furthermore, conceptus diapause may be a good paradigm to understand the interplay between the different genetic/molecular components of both the conceptus and endometrium at implantation.     

三带喙库蚊生殖滞育与其体内单糖成分变化的关系

本文报告用气相色谱法对乙型脑炎主要媒介三带喙库蚊上海株不同生殖状态蚊及不同季节采自野外蚊虫体内单糖及其变化的分析结果。色谱图中出现21个色谱峰,其中已知单糖8种,主要为半乳糖、葡萄糖与甘露糖等,其次为核糖、阿拉伯糖与鼠李糖等,不同生殖状态蚊的单糖组分及其含量具有一定的差异。实验室养殖蚊单糖组分及主要单糖含量比野外蚊多,野外蚊半乳糖、核糖与阿拉伯糖明显高于实验室养殖蚊。不同季节采获蚊的糖含量不同。繁殖蚊与解滞及夏季蚊核糖含量较高。短光照对核糖、鼠李糖等具有一定的抑制作用,但对半乳糖、葡萄糖的摄入与转化具有促进作用。     

天津地区主要空气污染物与胚胎停育关系的流行病学研究

目的初探天津地区主要空气污染物与孕早期胚胎停育之间的关系,从环境角度探讨影响胚胎生长发育、导致胚胎停育的可能危险因素。方法收集天津市市内六区15家综合性三甲医院和妇产科专科医院计划生育门诊2002年1月至2007年8月每月人工流产例数及当月胚胎停育病例的详细信息。获取同期天津市环境监测中心市内六区国控测点常规污染物（SO2、TSP、PM10、NO2）的月均浓度。以图表初探天津地区主要空气污染物与胚胎停育的关系,利用SPSS11．0统计软件分析各种污染物之间的相关性,不同月份、季节、采暖期与非采暖期胚胎停育占当月人工流产总数的百分比是否有差别。结果天津地区采暖期空气污染较非采暖期严重,且采暖期受孕者,其发生胚胎停育比例也相对较高,颗粒物质（PM10）、总悬浮颗粒物（TSP）的污染水平与胚胎停育的病例比例呈正相关。空气中的颗粒物质PM10可能是胚胎停育的危险因素,冬春季受孕可能对胚胎发育有潜在的负面影响。结论空气污染对人类胚胎的正常发育可能存在潜在的威胁。     

Cloning of cDNAs encoding Bombyx homologues of Cdc2 and Cdc2-related kinase from eggs

In diapausing eggs of the silkworm, Bombyx mori, embryonic cells are arrested at G₂ phase. The ability to undertake cell division is resumed in the course of diapause termination caused by such a treatment as acclimation to 5°C. As an initial trial to investigate the relationship between diapause and embryonic cell cycling, we have cloned and sequenced two Bombyx cDNAs encoding two distinct cdc2-related Ser/Thr protein kinases. One (Bm cdc2) encoded a 37.0 kDa protein which had all of the domains characteristic of other Cdc2 kinases. The other (Bcdrk) encoded a 45.1 kDa protein that was most similar to Drosophila and human cdc2-related protein kinases (Dcdrk protein and PISSRLE kinase). Northern blot analysis was carried out to examine levels of Bm cdc2 and Bcdrk mRNA during embryogenesis of non-diapause eggs. The result demonstrated that the mRNA level of Bm cdc2 appeared to correspond to the activity of nuclear/cellular division in non-diapause eggs, and that the developmental profile in the level of Bcdrk mRNA was somewhat different from that of Bm cdc2 mRNA.     

Autonomous regulation of the insect gut by circadian genes acting downstream of juvenile hormone signaling

In temperate regions, the shortening day length informs many insect species to prepare for winter by inducing diapause. The adult diapause of the linden bug, Pyrrhocoris apterus, involves a reproductive arrest accompanied by energy storage, reduction of metabolic needs, and preparation to withstand low temperatures. By contrast, nondiapause animals direct nutrient energy to muscle activity and reproduction. The photoperiod-dependent switch from diapause to reproduction is systemically transmitted throughout the organism by juvenile hormone (JH). Here, we show that, at the organ-autonomous level of the insect gut, the decision between reproduction and diapause relies on an interaction between JH signaling and circadian clock genes acting independently of the daily cycle. The JH receptor Methoprene-tolerant and the circadian proteins Clock and Cycle are all required in the gut to activate the Par domain protein 1 gene during reproduction and to simultaneously suppress a mammalian-type cryptochrome 2 gene that promotes the diapause program. A nonperiodic, organ-autonomous feedback between Par domain protein 1 and Cryptochrome 2 then orchestrates expression of downstream genes that mark the diapause vs. reproductive states of the gut. These results show that hormonal signaling through Methoprene-tolerant and circadian proteins controls gut-specific gene activity that is independent of circadian oscillations but differs between reproductive and diapausing animals.