Lamin B receptor regulates the growth and maturation of myeloid progenitors via its sterol reductase domain: implications for cholesterol biosynthesis in regulating myelopoiesis

Lamin B receptor (LBR) is a bifunctional nuclear membrane protein with N-terminal lamin B and chromatin-binding domains plus a C-terminal sterol Δ(14) reductase domain. LBR expression increases during neutrophil differentiation, and deficient expression disrupts neutrophil nuclear lobulation characteristic of Pelger-Hu?t anomaly. Thus, LBR plays a critical role in regulating myeloid differentiation, but how the two functional domains of LBR support this role is currently unclear. We previously identified abnormal proliferation and deficient functional maturation of promyelocytes (erythroid, myeloid, and lymphoid [EML]-derived promyelocytes) derived from EML-ic/ic cells, a myeloid model of ichthyosis (ic) bone marrow that lacks Lbr expression. In this study, we provide new evidence that cholesterol biosynthesis is important to myeloid cell growth and is supported by the sterol reductase domain of Lbr. Cholesterol biosynthesis inhibitors caused growth inhibition of EML cells that increased in EML-derived promyelocytes, whereas cells lacking Lbr exhibited complete growth arrest at both stages. Lipid production increased during wild-type neutrophil maturation, but ic/ic cells exhibited deficient levels of lipid and cholesterol production. Ectopic expression of a full-length Lbr in EML-ic/ic cells rescued both nuclear lobulation and growth arrest in cholesterol starvation conditions. Lipid production also was rescued, and a deficient respiratory burst was corrected. Expression of just the C-terminal sterol reductase domain of Lbr in ic/ic cells also improved each of these phenotypes. Our data support the conclusion that the sterol Δ(14) reductase domain of LBR plays a critical role in cholesterol biosynthesis and that this process is essential to both myeloid cell growth and functional maturation.     

Magi Is Associated with the Par Complex and Functions Antagonistically with Bazooka to Regulate the Apical Polarity Complex

The mammalian MAGI proteins play important roles in the maintenance of adherens and tight junctions. The MAGI family of proteins contains modular domains such as WW and PDZ domains necessary for scaffolding of membrane receptors and intracellular signaling components. Loss of MAGI leads to reduced junction stability while overexpression of MAGI can lead to increased adhesion and stabilization of epithelial morphology. However, how Magi regulates junction assembly in epithelia is largely unknown. We investigated the single Drosophila homologue of Magi to study the in vivo role of Magi in epithelial development. Magi is localized at the adherens junction and forms a complex with the polarity proteins, Par3/Bazooka and aPKC. We generated a Magi null mutant and found that Magi null mutants were viable with no detectable morphological defects even though the Magi protein is highly conserved with vertebrate Magi homologues. However, overexpression of Magi resulted in the displacement of Baz/Par3 and aPKC and lead to an increase in the level of PIP3. Interestingly, we found that Magi and Baz functioned in an antagonistic manner to regulate the localization of the apical polarity complex. Maintaining the balance between the level of Magi and Baz is an important determinant of the levels and localization of apical polarity complex.     

Structural biology of plasmodial proteins

Malaria is a global disease infecting several million individuals annually. Malarial infection is particularly severe in the poorest parts of the world and is a major drain on their limited resources. Development of drug resistance and absence of a preventive vaccine have led to an immediate necessity for identifying new drug targets to combat malaria. Understanding the intricacies of parasite biology is essential to design novel intervention strategies that can prevent the growth of the parasite. The structural biology approach towards this goal involves the identification of key differences in the structures of the human and parasite enzymes and the determination of unique protein structures essential for parasite survival. This review covers the work on structural biology of plasmodial proteins carried out during the period of January 2006 to June 2007.     

Modeling sequence evolution in acute HIV-1 infection

We describe a mathematical model and Monte Carlo (MC) simulation of viral evolution during acute infection. We consider both synchronous and asynchronous processes of viral infection of new target cells. The model enables an assessment of the expected sequence diversity in new HIV-1 infections originating from a single transmitted viral strain, estimation of the most recent common ancestor (MRCA) of the transmitted viral lineage, and estimation of the time to coalesce back to the MRCA. We also calculate the probability of the MRCA being the transmitted virus or an evolved variant. Excluding insertions and deletions, we assume HIV-1 evolves by base substitution without selection pressure during the earliest phase of HIV-1 infection prior to the immune response. Unlike phylogenetic methods that follow a lineage backwards to coalescence, we compare the observed data to a model of the diversification of a viral population forward in time. To illustrate the application of these methods, we provide detailed comparisons of the model and simulations results to 306 envelope sequences obtained from eight newly infected subjects at a single time point. The data from patients were in good agreement with model predictions, and hence compatible with a single-strain infection evolving under no selection pressure. The diversity of the samples from the other two patients was too great to be explained by the model, suggesting multiple HIV-1-strains were transmitted. The model can also be applied to longitudinal patient data to estimate within-host viral evolutionary parameters.     

Dynamic localization of tripartite motif-containing 22 in nuclear and nucleolar bodies

Tripartite motif-containing 22 (TRIM22) exhibits antiviral and growth inhibitory properties, but there has been no study on the localization and dynamics of the endogenous TRIM22 protein. We report here that TRIM22 is dramatically induced by progesterone in MDA-MB-231-derived ABC28 cells and T47D cells. This induction was associated with an increase in TRIM22 nuclear bodies (NB), and an even more prominent increase in nucleolar TRIM22 bodies. Distinct endogenous TRIM22 NB were also demonstrated in several other cell lines including MCF7 and HeLa cells. These TRIM22 NB resemble Cajal bodies, co-localized with these structures and co-immunoprecipitated with p80-coilin. However, IFNγ-induced TRIM22 in HeLa and MCF7 cells did not form NB, implying the forms and distribution of TRIM22 are regulated by specific cellular signals. This notion is also supported by the observation that TRIM22 NB undergoes dynamic cell-cycle dependent changes in distribution such that TRIM22 NB started to form in early G0/G1 but became dispersed in the S-phase. In light of its potential antiviral and antitumor properties, the findings here provide an interesting gateway to study the relationship between the different forms and functions of TRIM22.     

B30.2/SPRY domain in tripartite motif-containing 22 is essential for the formation of distinct nuclear bodies

Tripartite motif-containing 22 (TRIM22) is an important antiviral protein that forms distinct nuclear bodies (NB) in many cell types. This study aims to identify functional domains/residues for TRIM22’s nuclear localization and NB formation. Deletion of the really-interesting-new-gene (RING) domain, which is essential for its antiviral property, abolished TRIM22 NB formation. However, mutation of two critical residues Cys15 and Cys18 to alanine in the RING domain, did not affect NB formation notably. Although the deletion of the putative bipartite nuclear localization signal (NLS) abolished TRIM22 localization and NB formation, the B30.2/SplA and ryanodine receptor (SPRY) domain, and residues 491-494 specifically are also essential for nuclear localization and NB formation.     

Absence of CD34 on Murine Skeletal Muscle Satellite Cells Marks a Reversible State of Activation during Acute Injury

Background

Skeletal muscle satellite cells are myogenic progenitors that reside on myofiber surface beneath the basal lamina. In recent years satellite cells have been identified and isolated based on their expression of CD34, a sialomucin surface receptor traditionally used as a marker of hematopoietic stem cells. Interestingly, a minority of satellite cells lacking CD34 has been described.

Methodology/Principal Findings

In order to elucidate the relationship between CD34+ and CD34- satellite cells we utilized fluorescence-activated cell sorting (FACS) to isolate each population for molecular analysis, culture and transplantation studies. Here we show that unless used in combination with α7 integrin, CD34 alone is inadequate for purifying satellite cells. Furthermore, the absence of CD34 marks a reversible state of activation dependent on muscle injury.

Conclusions/Significance

Following acute injury CD34- cells become the major myogenic population whereas the percentage of CD34+ cells remains constant. In turn activated CD34- cells can reverse their activation to maintain the pool of CD34+ reserve cells. Such activation switching and maintenance of reserve pool suggests the satellite cell compartment is tightly regulated during muscle regeneration.