The Egr family of zinc finger transcription factors is rapidly induced by various mitogens and regulates cell growth, differentiation, and apoptosis. While it is clear that loss of Egr1 leads to anovulatory infertility due to LHβ deficiency in female mice, molecular function of Egr1 in male reproduction has not been clearly investigated. Here, we demonstrate that Egr1 acts as an intrinsic transcription factor in Leydig cells to regulate their proliferation and steroidogenesis in the testis as well as an extrinsic factor for male reproduction via LHβ transcription in the pituitary. Egr1 is predominantly expressed in spermatogonia and Leydig cells in immature testes and later detected in some of these cell types in mature testes. The fertility potential of Egr1(-/-) male mice is relatively deteriorated even at 2 month-old age and aggravated with aging. The incidence of abnormalities of seminiferous tubules such as Sertoli cell only was dramatically increased with aging. The number and mean size of Leydig cells were significantly reduced in Egr1(-/-) testes. The impairment of Leydig cells is consistent with significant reduction in levels of testosterone and expression of factors critical for steroidogenesis such as StAR in Egr1(-/-) testes. Exogenous administration of hCG rapidly and transiently induced Egr1 expression in Leydig cells culture in vitro. hCG could reinstate reduced mean size of Leydig cells but not reduced number of Leydig cells and aberrantly low StAR expression, suggesting that Egr1 has critical functions for Leydig cell proliferation and their steroidgenesis. In addition, daily sperm production and in vitro fertilization (IVF) competence were significantly reduced, and apoptosis was facilitated in these mice. Furthermore, hCG administration to compensate for relatively low LH levels in Egr1(-/-) males could not restore the compromised reproductive phenotypes such as IVF competence and apoptosis in these mice. Interestingly, expression of Egr2, a member of Egr family, is significantly elevated in Egr1(-/-) Leydig cells suggesting that genetic compensation of Egr2 may alleviate phenotypic aberration of Egr1(-/-) male testes. Collectively, these results suggest that Egr1 act as an intrinsic transcription factor required for proliferation and steroidogenesis of Leydig cells to govern spermatogenesis in the testis.

Testicular expression of CLDN11 (claudin-11), a tight junction protein was examined together with spermatogenesis and circulating testosterone levels in Korean soft-shelled turtle (Pelodiscus maackii). Spermatogenesis started during the breeding season in May and peaked in August when the breeding season ended. Spermiation started in July and peaked in October, showing the typical pattern of spermatogenesis in temperate zone reptiles. Deduced amino acid sequences of P. maackii CLDN11 was highly homologous to those of avian and mammals, suggesting the conserved nature of CLDN11 in amniotes. During the non-breeding season when the spermatogenesis was active and circulating testosterone levels elevated, testicular CLDN11 mRNA and protein (19kDa) levels were high. Strong, wavy CLDN11 immunoreactive strands run parallel to basement membrane in the basal part of the seminiferous epithelium, delaminating the spermatogonia and early spermatocytes in the open compartment. Otherwise, CLDN11 was found beneath the early spermatocytes and in the Sertoli cell cytoplasm perpendicular to basement membrane. In double labeling experiment, punctate ZO-1 immunoreactivity was found within the CLDN11 strands run parallel to the basement membrane as well as at the most periphery of seminiferous epithelium where ZO-1 and CLDN11 in Sertoli cells were mostly cytoplasmic and perpendicular to basement membrane. Together, recruit of CLDN11 and ZO-1 to the inter-Sertoli TJs was tightly coupled with spermatogenic stage. At the breeding season when the circulating testosterone levels and spermatogenic activity remained low, testicular CLDN11 mRNA and protein levels were low. CLDN11 was found at apicolateral contacts between adjacent Sertoli cells devoid of the postmeiotic germ cells, suggesting that CLDN11 between adjacent Sertoli cells also participates in the maintenance of seminiferous lumen. In P. maackii testis, CLDN11 as a structural element of the blood-testis barrier dynamically changed according to spermatogenic activity and circulating androgen levels. This is the first study on the CLDN TJs at the BTB in reptilian testis.

This study is a basic study on the development of functional substances involved in obesity prevention, lipid metabolism, and immune regulation. Male Sprague-Dawley rats were fed a high-fat diet for 10 weeks. Allium monanthum extracts (AME) were administered orally to obesity-induced rats, and their lipid-lowering, antioxidative and various types of biological effects related to the immune system were examined. Blood free fatty acid and triglyceride concentrations decreased as the dose of AME increased. Total cholesterol and LDL cholesterol concentrations in the blood decreased as the dose of AME increased. The total cholesterol concentrations in the liver of the AME-treated groups were lower than the control group. The thiobarbituric acid reactive concentrations were lower in the plasma and liver of all AME-treated groups than the control group. Plasma AST and ALT activities did not show any significant differences among the treatment groups. IL-1β and IL-6 concentrations in the liver tended to decrease as the dose of AME increased. TNF-α and IL-10 concentrations did now show any significant differences compared to the control group. Lower expression levels of TNF-α, Apo-B and Apo-E genes were found in the AME-treated groups. Taken together, these results indicate that AME may show positive effects in lipid lowering, antioxidation and anti-inflammation.

Aquaporin 5 (AQP5) implicated in the generation of saliva, tears, and pulmonary secretions functions as a water-specific channel. Epididymal epithelial cells actively reabsorb water, ions and proteins. Large quantity of testicular fluid movement across the epididymal tubules generates high osmotic milieu which is important for sperm maturation. In an effort to understand the fluid homeostasis and its regulation by sex steroids in male reproductive tract, the expression of AQP5 was examined in different regions of mouse epididymis during postnatal development. The effect of androgen on the expression of epididymal AQP5 was examined in ORX model. AQP5 mRNA levels were the highest in corpus region in which drastic increase was noted during sexual maturation. Epididymal AQP5 immunoreactivity was largely found in apical as well as basal region of luminal epithelia. Moderate immunoreactivity for AQP5 was found in in smooth muscle cells in both immature and mature mice. Epididymal lumen of ORX mice showed shrinkage together with decrease in AQP5 expression. Alteration of AQP5 expression in ORX epididymis was partially recovered by androgen injection. AQP5 mRNA was induced at 10uM 5α-DHT in organ cultured epididymis. Chromatin immunoprecipitatin (ChIP) showed that 5α-DHT induced recruitment of androgen receptor (AR) to the －4635 to －4453 bp region of the AQP5 gene promoter in adult epididymis. Taken together, axial regulatory mechanism may control transcription of AQP5 along the length of epididymal tubule. Water transport through AQP5 is important for late sperm maturation and storage in epididymis. Androgen may directly induce AQP5 gene transcription via activation of AR in epididymis.

Foxi1, a forkhead family of transcription factor, in narrow and clear cells in epididymis is required for male fertility through regulating transcription of vacuolar H+-ATPase. To understand the regulation of Foxi1 gene activation in epididymis, the effects of steroids and their receptor antagonists and testicular factors on the expression of Foxi1 in epididymal segments were examined in mouse. Epididymis were sampled from adult mice following injections of ICI 182,780 (5mg/head, 2 times for 15 days), dexamethasone (DEX, 0.1,1,10ug/kg/day for 5 days) or oral administration of flutamide (FLM, 100mg/kg/day for 10 days). Otherwise, adult mice were orchidectomized (ORX), rested for 2 weeks, and received testosterone propionate(TP, 3mg/kg/day) for 7 days. In addition, adult male mice were subjected to efferent duct ligation (EDL) and epididymis was collected after 15 days. To study estrogen regulation of Foxi1 gene activation via estrogen receptor α (ESR1), Foxi1 expression was examined in ESR1 knock-out mice epididymis. Expression and subcellular localization of Foxi1 was analyzed by realtime RT-PCR and immunohistochemistry. To search transcription factor binding in the mouse Foxi1 gene promoter, in silico analysis was performed using TESS, TFSEARCH, and Gene-Regulation. ICI 182,780 significantly decreased Foxi1 mRNA levels in caput and corpus but increased in cauda epididymis. Foxi1 mRNA levels in caput epididymis of ESR1 KO mice were significantly lower than those of WT mice, but no significantly changed in corpus and cauda epididymis. Taken together, estrogen differentially regulates Foxi1 gene expression in epididymis. In ORX mice, Foxi1 mRNA levels were significantly increased in epididymis, and which was abrogated by TP. Though FLM did not significantly alter the Foxi1 mRNA levels, androgen may affect Foxi1 gene expression in epididymis. DEX significantly decreased Foxi1 mRNA levels in caput and corpus epididymis at 0.1ug/kg/day and in cauda epididymis at 1ug/kg/day, suggesting that glucocorticoid may negatively regulate Foxi1 gene expression. No significant change in Foxi1 mRNA levels was found after EDL. Foxi1 immunoreactivity was found in the nuclei of narrow cells of caput epididymis including initial segment and clear cells of corpus and cauda epididymis. Of note, in ORX mice, Foxi1-positive narrow cells and clear cells were increased, and which was abrogated by TP. In silico analysis revealed the presence of putative binding sequences for ESR1, AR, and GR in the 5’ upstream region from the Foxi1 promoter. In conclusion, the expression of Foxi1 in narrow cells in caput epididymis might be positively regulated by estrogen via ESR1, which was different from estrogen–ESR signaling in clear cells in corpus and cauda epdididymis. Androgen and glucocorticoid may negatively regulate expression of Foxi1 in all epdididymial segments.

1. 본 연구에서는 완두유래의 세포질성 PsAPX1 유전자를 대상으로 산화스트레스 유도성 프로모터를 연결하여 엽록체에 targeting 되는 과발현 운반체를 제작하고 벼에 도입한 결과 형질전환체에서 도입유전자 수가 1~3 copy인 것으로 나타나, 적은 수의 유전자가 안정적으로 도입되었음을 확인하였다. 2. 염, 오존, 자외선, 한발과 같은 다양한 환경스트레스 조건에서 내성이 증진된 우수 계통을 선발하기 위하여 작성된 형질전환 벼 계통들을 대상으로 생물

This experiment was conducted to investigate the changes of growth and maturity and to clarify the function of supernodulating characters, excessive nodules and high biological nitrogen fixation rate (BNF), on maturity in response to different planting time in supernodulating soybean mutants. Two supernodulating soybean mutants, Sakukei4 and SS2-2, and their parent cultivars, Enrei and Shinpaldalkong2, were planted on May 24 and June 15, 2004. The degrees of the shortening of growth days by the planting time delay were 18 to 22 days in four cultivar, and there were no significant differences among the cultivars. However, four cultivars showed the different maturity properties. Sakukei4, mutated from Enrei, showed later maturity than that of Enrei, and 882-2, mutated from Shinpaldalkong2, showed earlier maturity than that of Shinpaldalkong2. The plant and nodule dry weights at R6 stage of Sakukei4 showed the smallest decrement and those of SS2-2 was showed the largest decrement by the delay of planting time. The photosynthetic rates of Sakukei4 during the late reproductive growth period were slowly decreased, however those of SS2-2 were steeply decreased in two planting time treatments. Overall, the growth of Sakukei4 was decreased slowly, however the growth of SS2-2 was decreased sharply according to the delay of planting time. The percentage of seed yield of Sakukei4 in June planting plot compared with May planting plot at R8 stage was 92~% , which was the lowest decreasing rate of yield among the cultivars, and in the case of SS2-2, it was in 76~% , the highest one. These results indicated that the responses of supernodulating mutants by the delay of planting time were very similar to the wild types. This means supernodulating characters in supernodulating soybean mutants might not affect to the maturity property. Additionally, the maturity property could be considered as an important characteristics to decide or to select on the developments of supernodulating soybean mutants, which have a low productivity by an excessive nodules, especially.

This experiment was conducted to clarify the functions of supernodulating characters on seed yield determination through the comparison of agricultural traits of supernodulating soybean mutants, Sakukei4, SS2-2, and their parent cultivars, Enrei and Shinpaldalkong2. The plant dry weights of supernodulating mutants, Sakukei4 and SS2-2, were 52~% and 61~% of their wild type parents at full seed stage (R6). However, the relative growth rate (RGR) from the pod set stage (R3) to R6 of Sakukei4 was 0.022 g/g/day and that of SS2-2 was 0.016 g/g/day, which were higher than those of their parents. Nodule number and dry weight were increased in two supernodulating mutants by the R6 stage. The nitrogen concentrations of leaf, petiole and stem of Sakukei4 were higher than those of Enrei. SS2-2 showed higher nitrogen concentration in petiole than Shinpaldalkong2 had. The positive correlations were appeared between nodule dry weight, plant dry weight and pod number, in two supernodulating mutants during the period from R3 to R6 stage. Although all of the yield components and seed yield were lower in two supernodulating mutants than their parents at the stage of full maturity (R8), the harvest index was higher in supernodulating mutants. The increasing rates of pod number to stem dry weight in two supernodulating mutants showed the higher than those of two their parents at R8 stage. In conclusion, the relative growth rates during the early to the middle reproductive growth period were higher in supernodulating mutants than the wild types. This could be resulted in an increase in pod number. The increase of relative growth rate was the result of the successive supplement of nitrogen source from biological nitrogen fixation (BNF) of nodules during the middle reproductive growth period in supernodulating mutants.

This experiment was conducted to investigate the soil nitrogen credit of biological nitrogen fixation (BNF) and the nitrogen balance of soybean in soybean-barley cropping systems. Soybean cultivar, Shinpaldalkong2 and barley cultivar, Olbori, were used in soybean mono-cropping (SM), barley monocropping (BM), and barley­soybean double cropping system. The barley-soybean double cropping system was treated with two different levels of nitrogen fertilizers, 0 nitrogen fertilizer (BS-F0), and standard nitrogen fertilizer (BS-F1). Nitrogen and organic matter concentrations in soil of BS-F1 plot on October, 2001 were increased 4.8~%~;and~;5.9~% , respectively, compared with those on October, 2000. The ranges of BNF rate in soybean were 69.1~~ 88.2~% in two years, and the rate was the highest in BS-F0 plot and the lowest in SM plot. The ranges of nitrogen harvest index (NHI) in all treatments were 83.9~~86.7~% . The yield was 270 kg/10a in BS­F1 plot and 215 kg/10a in BS-F0 plot. However, the nitrogen balances were +0.6 kg/10a of gain of soil nitrogen in BS-F0 plot and -0.4 kg/10a of loss of soil nitrogen in BS-F1 plot. In comparisons of SM and BS-F1 plots, although the seed yields were similar in two plots, the loss of soil nitrogen was higher in SM than BS-F1 plot. Overall, our results suggest that barley-soybean double cropping system was more effective in respect to seed productivity and soil nitrogen conservation than soybean monocropping system, and the N credit to following crops by soybean cultivation was identified in soybean double cropping system.

The diagnostic software for the wastewater treatment plant using activated-sluge process is developed in order to increase the efficiency of management of the wastewater treatment plant. This software is based on the expert system and the visualized user interface including the diagnosis of quantitative and qualitative data. For the generalization of this software, the initialization of each unit process and updating the files can be possible.