Preparation and Activity Identification of Recombinant Protein PACAP-PTD

[Objective] This study aimed to prepare recombinant protein PACAP-PTD and measure its activity. [Method] The gene that encodes fusion protein PACAP-PTD was cloned into the expression vector pKYB to construct recombinant expression vector pKYB-PACAP-PTD, which was then transformed into E. coli ER2566. The fusion protein consisting of PACAP-PTD, intein and chitin was expressed under the induction of IPTG. Finally, the target fusion protein PACAP-PTD was purified with IMPACT system （ Intein Mediated Purification with an Affinity of chitin-binding Tag）, and its activities to cross blood-brain barrier and to promote cell proliferation were measured. [ Result~ The molecular weight of the fusion protein PACAP-PTD determined with laser time-of-flight mass spectrometry was con- sistent with the theoretical value. In addition, the protein could effectively cross the blood-brain barrier and promote cell proliferation as well. [ Conclusion] The construction and preparation of the fusion protein PACAP-PTD not only lays foundation for further study on its biological function, but also improves the route of PACAP administration, and thus expands its scope of application.

Reversal of idiopathic hypogonadotropic hypogonadism： a cohort study in Chinese patients

Although idiopathic hypogonadotropic hypogonadism （IHH） has traditionally been viewed as a life-long disease caused by a deficiency of gonadotropin-releasing hormone neurons, a portion of patients may gradually regain normal reproductive axis function during hormonal replacement therapy. The predictive factors for potential IHH reversal are largely unknown. The aim of our study was to investigate the incidence and clinical features of IHH male patients who had reversed reproductive axis function. In this retrospective cohort study, male IHH patients were classified into a reversal group （n = 18） and a nonreversal group （n = 336）. Concentration of gonadotropins and testosterone, as well as testicle sizes and sperm counts, were determined. Of 354 IHH patients, 18 （5.1%） acquired normal reproductive function during treatment. The median age for reversal was 24 years old （range 21-34 years）. Compared with the nonreversal group, the reversible group had higher basal luteinizing hormone （LH） （1,0±0.7 IU 1-1 vs 0.4±0.4 IU 1-1, P 〈 0.05） and stimulated LH （28.3 ± 22.6 IU 1-1 vs 1.9 ±1.1 IU 1-1, P 〈 0.01） levels, as well as larger testicle size （5.1 ±2.6 ml vs 1.5± 0.3 ml, P〈 0.01）, at the initial visit. In summary, larger testicle size and higher stimulated LH concentrations are favorite parameters for reversal. Our finding suggests that reversible patients may retain partially active reproductive axis function at initial diagnosis.

Fast-Suppressor Screening for New Components in Protein Trafficking,Organelle Biogenesis and Silencing Pathway in Arabidopsis thaliana Using DEX-Inducible FREE1-RNAi Plants

Membrane trafficking is essential for plant growth and responses to external signals.The plant unique FYVE domain-containing protein FREE1 is a component of the ESCRT complex（endosomal sorting complex required for transport）.FREE1 plays multiple roles in regulating protein trafficking and organelle biogenesis including the formation of intraluminal vesicles of multivesicular body（MVB）,vacuolar protein transport and vacuole biogenesis,and autophagic degradation.FREE1 knockout plants show defective MVB formation,abnormal vacuolar transport,fragmented vacuoles,accumulated autophagosomes,and seedling lethality.To further uncover the underlying mechanisms of FREE1 function in plants,we performed a forward genetic screen for mutants that suppressed the seedling lethal phenotype of FREE1-RNAi transgenic plants.The obtained mutants are termed as suppressors of free1（sof）.To date,229 putative sof mutants have been identified.Barely detecting of FREE1 protein with M3 plants further identified 84 FREE1-related suppressors.Also145 mutants showing no reduction of FREE1 protein were termed as RNAi-related mutants.Through next-generation sequencing（NGS）of bulked DNA from F2 mapping population of two RNAi-related sof mutants,FREE1-RNAi T-DNA inserted on chromosome 1 was identified and the causal mutation of putative sof mutant is being identified similarly.These FREE1-and RNAi-related sof mutants will be useful tools and resources for illustrating the underlying mechanisms of FREE1 function in intracellular trafficking and organelle biogenesis,as well as for uncovering the new components involved in the regulation of silencing pathways in plants.

Evolutionary characteristics of exons expressed at different abundance levels in mammals

With comparative genomics approaches, we evaluated the evolutionary characteristics of conservation of exons which are expressed abundantly, moderately or lowly in mammals. Using non-coding regions and pseudogenes as controls, sequence identity, phastCons and Ka/Ks analyses were carried out and our results showed that as the exons of high abundance are highly conserved, the minor and low exons also showed conservative characteristics in evolution. Our findings suggested that the exons with less abundance which constitute a large proportion of distinct species in transcriptome of organisms are under functional constraint and might play certain roles in enriching biological complexity in the evolution of organisms.

DNA Copy Number Aberrations in Breast Cancer by Array Comparative Genomic Hybridization

Array comparative genomic hybridization （CGH） has been popularly used for analyzing DNA copy number variations in diseases like cancer. In this study, we investigated 82 sporadic samples from 49 breast cancer patients using 1-Mb resolution bacterial artificial chromosome （2GH arrays. A number of highly frequent genomic aberrations were discovered, which may act as ＂drivers＂ of tumor progression. Meanwhile, the genomic profiles of four ＂normal＂ breast tissue samples taken at least 2 cm away from the primary tumor sites were also found to have some genomic aberrations that recurred with high frequency in the primary tumors, which may have important implications for clinical therapy. Additionally, we performed class comparison and class prediction for various clinicopathological parameters, and a list of characteristic genomic aberrations associated with different clinicopathological phenotypes was compiled. Our study provides clues for further investigations of the underlying mechanisms of breast carcinogenesis.

3D Genome Tuner:Compare Multiple Circular Genomes in a 3D Context

Circular genomes, being the largest proportion of sequenced genomes, play an important role in genome analysis. However, traditional 2D circular map only provides an overview and annotations of genome but does not offer feature-based comparison. For remedying these shortcomings, we developed 3D Genome Tuner,a hybrid of circular map and comparative map tools. Its capability of viewing comparisons between multiple circular maps in a 3D space offers great benefits to the study of comparative genomics. The program is freely available （under an LGPL licence） at http：//sourceforge.net/projects/dgenometuner.

Evolutionary Transients in the Rice Transcriptome

In the canonical version of evolution by gene duplication, one copy is kept unaltered while the other is free to evolve. This process of evolutionary experimentation can persist for millions of years. Since it is so short lived in comparison to the lifetime of the core genes that make up the majority of most genomes, a substantial fraction of the genome and the transcriptome may—in principle—be attributable to what we will refer to as ＂evolutionary transients＂, referring here to both the process and the genes that have gone or are undergoing this process. Using the rice gene set as a test case, we argue that this phenomenon goes a long way towards explaining why there are so many more rice genes than Arabidopsis genes, and why most excess rice genes show low similarity to eudicots.