Differential gene dosage effects of Ad4BP/SF-1 on target tissue development

Ad4BP/SF-1 (NR5A1) was identified as a key regulator of the hypothalamus-pituitary-gonadal and -adrenal axes. Loss-of-function studies revealed that Ad4BP/SF-1 is essential for the development of these tissues and spleen. Here, we generated transgenic mouse with BAC recombinants carrying a dual promoter and Tet-off system. These recombinants have a potential to express lacZ and Ad4BP/SF-1 in the tissues where endogenous Ad4BP/SF-1 is expressed. However, protein level of Ad4BP/SF-1 varied among the tissues of the transgenic mice and probably thereby the target tissues are affected differentially. The BAC-transgenic mice were applied to rescue Ad4BP/SF-1 KO mouse. Interestingly, the mice successfully rescued the gonad and spleen but failed to rescue the adrenal gland. This variation might be dependent on in part the protein expression levels among the tissues and in part on differential sensitivities to the gene dosage.     

Cdc2-like kinase 2 (Clk2) promotes early neural development in Xenopus embryos

Neural induction and patterning in vertebrates are regulated during early development by several morphogens, such as bone morphogenetic proteins (BMPs) and fibroblast growth factors (FGFs). Ventral ectoderm differentiates into epidermis in response to BMPs, whereas BMP signaling is tightly inhibited in the dorsal ectoderm which develops into neural tissues. Here, we show that Cdc2-like kinase 2 (Clk2) promotes early neural development and inhibits epidermis differentiation in Xenopus embryos. clk2 is specifically expressed in neural tissues along the anterior-posterior axis during early Xenopus embryogenesis. When overexpressed in ectodermal explants, Clk2 induces the expression of both anterior and posterior neural marker genes. In agreement with this observation, overexpression of Clk2 in whole embryos expands the neural plate at the expense of epidermal ectoderm. Interestingly, the neural-inducing activity of Clk2 is increased following BMP inhibition and activation of the FGF signaling pathway in ectodermal explants. Clk2 also downregulates the level of p-Smad1/5/8 in cooperation with BMP inhibition, in addition to increasing the level of activated MAPK together with FGF. These results suggest that Clk2 plays a role in early neural development of Xenopus possibly via modulation of morphogen signals such as the BMP and FGF pathways.     

Reducing IRS-1 Activation Cause Mutation of Tyrosine Kinase Domain hINSR Gene on Type-2 Diabetes Mellitus Patients

The purpose of this study is to examine the effect of mutation on tyrosine kinase hINSR gene of DM Type 2 patients reduce the IRS- 1 activation by in silico analysis. Blood DNA of DM Type 2 patients from Saeful Anwar Hospital Malang were amplified and sequenced by specific primers of tyrosine kinase domain of hINSR gene. These gene sequences were converted to protein sequence by BLAST and the IRS-1 protein sequence is retrieved from NCBI database. Both of the protein sequence was aligned by using Bio edit version 5.0.6. The model of three dimension protein was predicted by SWISS MODEL webserver, and visualized the structure alteration by using Pymol 0.99rc6 and Hex 5.0, and then superimpose of the hINSR and IRS-1 interaction were examined by docking using Hex 5.0. The results showed that one substitution and one deletion of 8-3F patient exon-22 hINSR gene tyrosine kinase domain cause loss of four helixes and three coils structures on tyrosine kinase hINSR protein. Six-deletions and six-substitutions on same gene domain of DMK9 patient changed the two helixes became coil structure. The binding energy of hINSR tyrosine kinase with IRS-1 of normal is E= -494.67 kJ/mol, DMK9 patient is E= -458.4 kJ/mol, and 8-3F patient is E=-544.20 kJ/mol. The DMK9 patient prognosis has better physiological condition than 8-3F patient. Interaction between 8-3F of hINSR tyrosine kinase domain mutation and PTB domain IRS-1 is more spontaneous than DMK9, but both of them were reduced on IRS-1 activation respectively.