Exportin 4 Interacts with Sox9 through the HMG Box and Inhibits the DNA Binding of Sox9

Sox9 is a transcription factor that is required for tissue development in mammals. In general, such transcription factors require co-regulators for precise temporal and spatial control of the activation and inactivation of the numerous genes necessary for precise development during embryogenesis. Here we identify a new Sox9 co-regulator: Using affinity chromatography with immobilized Sox9 protein, we identified exportin 4 (Exp4) as an interacting protein of Sox9 in human cultured cells. Interaction between endogenous Exp4 and Sox9 proteins was confirmed in the human osteosarcoma U2OS cells by immunoprecipitation experiments using anti-Sox9 antibody. siRNA depletion of Exp4 enhanced transcription of Sox9 target genes in U2OS cells, but did not affect nuclear localization of Sox9. These results suggest that Exp4 regulates Sox9 activity in the nucleus. Furthermore we found that the HMG box of Sox9 was responsible for binding to Exp4, and the HMG box was required for suppression of Sox9-mediated transcription. This contrasts with the known Sox9 co-regulators which bind to its transcriptional activation domain. Chromatin immunoprecipitation analyses revealed that Exp4 prevents Sox9 binding to the enhancers of its target genes. These results demonstrate that Exp4 acts as a Sox9 co-regulator that directly regulates binding of Sox9 to its target genes

A SRY-HMG box frame shift mutation inherited from a mosaic father with a mild form of testicular dysgenesis syndrome in Turner syndrome patient

Background: Sex determining factor (SRY) located on the short arm of the Y chromosome, plays an important role in initiating male sex determination, resulting in development of testicular tissue. Presence of the SRY gene in females results in XY sex reversal and increased risk of gonadal germ cell tumours if the karyotype also includes the so-called GonadoBlastoma on the Y chromosome (GBY) region. The majority of mutations within the SRY gene are de novo affecting only a single individual in the family. The mutations within the high-mobility group (HMG) region have the potential to affect its DNA binding activity.Case Presentation: We performed G- and R-banding cytogenetic analysis of the patient and her family members including her father. We also performed molecular genetic analysis of SRY gene. Cytogenetic analysis in the patient (Turner Syndrome) revealed the mosaic karyotype as 45, X/46, XY (79%/21% respectively) while her father (milder features with testicular dysgenesis syndrome) has a normal male karyotype (46, XY). Using molecular approach, we screened the patient and her father for mutations in the SRY gene. Both patient and her father showed the same deletion of cytosine within HMG box resulting in frame shift mutation (L94fsX180), the father in a mosaic pattern. Histological examination of the gonads from the patient revealed the presence of gonadoblastoma formation, while the father presented with oligoasthenozoospermia and a testicular seminoma. The frameshift mutation at this codon is novel, and may result in a mutated SRY protein.Conclusion: Our results suggest that lack of a second sex chromosome in majority cells of the patient may have triggered the short stature and primary infertility, and the mutated SRY protein may be associated with the development of gonadoblastoma. It is of importance to note that mosaic patients without a SRY mutation also have a risk for malignant germ cell tumors

Localization and potential role of matrix metalloproteinase-1 and tissue inhibitors of metalloproteinase-1 and -2 in different phases of bronchopulmonary dysplasia

Bronchopulmonary dysplasia (BPD) can evolve in prematurely born infants who require mechanical ventilation because of hyaline membrane disease (HMD). The development of BPD can be divided in an acute, a regenerative, a transitional, and a chronic phase. During these different phases, extensive remodeling of the lung parenchyma with re-epithelialization of the alveoli and formation of fibrosis occurs. Matrix metalloproteinase-1 (MMP-1) is an enzyme that is involved in re-epithelialization processes, and dysregulation of MMP-1 activity contributes to fibrosis. Localization of MMP-1 and its inhibitors, tissue inhibitor of metalloproteinase (TIMP)-1 and TIMP-2, were investigated in lung tissue obtained from infants who died during different phases of BPD development. In all studied cases (n = 50) type-II pneumocytes were found to be immunoreactive for MMP-1, TIMP-1, and TIMP-2. During the acute and regenerative phase of BPD, type-II pneumocytes re-epithelialize the injured alveoli. This may suggest that MMP-1 and its inhibitors, expressed by type-II pneumocytes, play a role in the re-epithelialization process after acute lung injury. Although MMP-1 staining intensity remained constant in type-II pneumocytes during BPD development, TIMP-1 increased during the chronic fibrotic phase. This relative elevation of TIMP-1 compared with MMP-1 is indicative for reduced collagenolytic activity by type-II pneumocytes in chronic BPD and may contribute to fibrosis. Fibrotic foci in chronic BPD contained fibroblasts immunoreactive for MMP-1 and TIMP-1 and -2. This may indicate that decreased collagen turnover by fibroblasts contributes to fibrosis in BPD development

Cytosolic SYT/SS18 Isoforms Are Actin-Associated Proteins that Function in Matrix-Specific Adhesion

SYT (SYnovial sarcoma Translocated gene or SS18) is widely produced as two isoforms, SYT/L and SYT/S, that are thought to function in the nucleus as transcriptional coactivators. Using isoform-specific antibodies, we detected a sizable pool of SYT isoforms in the cytosol where the proteins were organized into filamentous arrays. Actin and actin-associated proteins co-immunoprecipitated with SYT isoforms, which also co-sedimented and co-localized with the actin cytoskeleton in cultured cells and tissues. The association of SYT with actin bundles was extensive yet stopped short of the distal ends at focal adhesions. Disruption of the actin cytoskeleton also led to a breakdown of the filamentous organization of SYT isoforms in the cytosol. RNAi ablation of SYT/L alone or both isoforms markedly impaired formation of stress fibers and focal adhesions but did not affect formation of cortical actin bundles. Furthermore, ablation of SYT led to markedly impaired adhesion and spreading on fibronectin and laminin-111 but not on collagen types I or IV. These findings indicate that cytoplasmic SYT isoforms interact with actin filaments and function in the ability cells to bind and react to specific extracellular matrices

Search for anisotropic gravitational-wave backgrounds using data from Advanced LIGO and Advanced Virgo's first three observing runs

We report results from searches for anisotropic stochastic gravitational-wave backgrounds using data from the first three observing runs of the Advanced LIGO and Advanced Virgo detectors. For the first time, we include Virgo data in our analysis and run our search with a new efficient pipeline called {\tt PyStoch} on data folded over one sidereal day. We use gravitational-wave radiometry (broadband and narrow band) to produce sky maps of stochastic gravitational-wave backgrounds and to search for gravitational waves from point sources. A spherical harmonic decomposition method is employed to look for gravitational-wave emission from spatially-extended sources. Neither technique found evidence of gravitational-wave signals. Hence we derive 95\% confidence-level upper limit sky maps on the gravitational-wave energy flux from broadband point sources, ranging from $F_{\alpha, \Theta} < {\rm (0.013 - 7.6)} \times 10^{-8} {\rm erg \, cm^{-2} \, s^{-1} \, Hz^{-1}},$ and on the (normalized) gravitational-wave energy density spectrum from extended sources, ranging from $\Omega_{\alpha, \Theta} < {\rm (0.57 - 9.3)} \times 10^{-9} \, {\rm sr^{-1}}$, depending on direction ($\Theta$) and spectral index ($\alpha$). These limits improve upon previous limits by factors of $2.9 - 3.5$. We also set 95\% confidence level upper limits on the frequency-dependent strain amplitudes of quasimonochromatic gravitational waves coming from three interesting targets, Scorpius X-1, SN 1987A and the Galactic Center, with best upper limits range from $h_0 < {\rm (1.7-2.1)} \times 10^{-25},$ a factor of $\geq 2.0$ improvement compared to previous stochastic radiometer searches.Comment: 23 Pages, 9 Figure

Diving below the spin-down limit:constraints on gravitational waves from the energetic young pulsar PSR J0537-6910

We present a search for continuous gravitational-wave signals from the young, energetic X-ray pulsar PSR J0537-6910 using data from the second and third observing runs of LIGO and Virgo. The search is enabled by a contemporaneous timing ephemeris obtained using NICER data. The NICER ephemeris has also been extended through 2020 October and includes three new glitches. PSR J0537-6910 has the largest spin-down luminosity of any pulsar and is highly active with regards to glitches. Analyses of its long-term and inter-glitch braking indices provided intriguing evidence that its spin-down energy budget may include gravitational-wave emission from a time-varying mass quadrupole moment. Its 62 Hz rotation frequency also puts its possible gravitational-wave emission in the most sensitive band of LIGO/Virgo detectors. Motivated by these considerations, we search for gravitational-wave emission at both once and twice the rotation frequency. We find no signal, however, and report our upper limits. Assuming a rigidly rotating triaxial star, our constraints reach below the gravitational-wave spin-down limit for this star for the first time by more than a factor of two and limit gravitational waves from the l = m = 2 mode to account for less than 14% of the spin-down energy budget. The fiducial equatorial ellipticity is limited to less than about 3 x 10⁻⁵, which is the third best constraint for any young pulsar