Role of Gbx2 expression in cranial nerve V development

Abstract only availableGbx2 is a member of the Gbx family of homeobox genes that encodes for the transcriptional factors Gbx1 and Gbx2. The amino acid sequence of Gbx2 is highly conserved across multiple species (e.g. mice, zebrafish, chicken, and frogs) with nearly 100% sequence identity between the Gbx2 homeodomains of the aforementioned species. Expression studies have shown that Gbx2 is expressed early during gastrulation in the posterior neural plate (Bouillet et al., 1995) and prospective anterior hindbrain (Niss and Leutz, 1998). The function of Gbx2 in mice and zebrafish has been extensively studied with loss-of-function as well as hypomorphic models in both species. Results of these studies have shown that the Gbx2 homeobox gene is required for normal development of the isthmic organizer at the midbrain hindbrain boundary (MHB) that is responsible for patterning the midbrain and the anterior hindbrain. The hindbrain controls many basic life functions such as breathing and heartbeat. In early vertebrate development, the hindbrain is partitioned into seven or eight distinct segments called rhombomeres (r). Rhombomeres give rise to differentiated brain regions such as the cerebellum, pons, and medulla oblongata. Previous studies had shown that cranial nerve V (nV), derived from r2 and r3, fails to develop normally in Gbx2 hypomorphic mice. The nV motor axons innervate several muscles, including those in the jaw required to suckle and masticate. These Gbx2 hypomorphic mice die immediately after birth presumably due to an inability to nurse on their mother as wild-type mice do. Our preliminary studies have shown that zebrafish Gbx2 also affects nV development. Our research this summer was focused around examining if mouse Gbx2 can rescue the abnormalities in nV caused by injecting morpholino. To accomplish this, we will attempt to rescue the hindbrain phenotype in zebrafish embryos by simultaneously injecting zebrafish morpholino with synthesized mouse Gbx2 mRNA.Life Sciences Undergraduate Research Opportunity Progra

Novel Approaches to Improve Myeloma Cell Killing by Monoclonal Antibodies

The monoclonal antibodies (mAbs) have significantly changed the treatment of multiple myeloma (MM) patients. However, despite their introduction, MM remains an incurable disease. The mAbs currently used for MM treatment were developed with different mechanisms of action able to target antigens, such as cluster of differentiation 38 (CD38) and SLAM family member 7 (SLAMF7) expressed by both, MM cells and the immune microenvironment cells. In this review, we focused on the mechanisms of action of the main mAbs approved for the therapy of MM, and on the possible novel approaches to improve MM cell killing by mAbs. Actually, the combination of anti-CD38 or anti-SLAMF7 mAbs with the immunomodulatory drugs significantly improved the clinical effect in MM patients. On the other hand, pre-clinical evidence indicates that different approaches may increase the efficacy of mAbs. The use of trans-retinoic acid, the cyclophosphamide or the combination of anti-CD47 and anti-CD137 mAbs have given the rationale to design these types of combinations therapies in MM patients in the future. In conclusion, a better understanding of the mechanism of action of the mAbs will allow us to develop novel therapeutic approaches to improve their response rate and to overcome their resistance in MM patients

Identification of <i>PSMB4</i> and <i>PSMD4</i> as novel target genes correlated with 1q21 amplification in patients with smoldering myeloma and multiple myeloma

Multiple myeloma (MM) is a malignant plasma cell (PC) dyscrasia characterized by heterogeneous biological features and genetic alterations, resulting in a wide range of disease courses.1,2 Despite all the therapeutic strategies developed in the last three decades, MM is still incurable, and almost all patients will inevitably experience disease progression and eventually relapse.3 Among all the genetic abnormalities, the amplification of the 1q21 region is one of the most frequent cytogenetic abnormalities occurring in malignant PC and it has become a new prognostic factor in MM patients.4,5 The incidence of gain and/or amplification of the 1q21 locus (1q21+) increases with disease progression. It can be detected in around 30-45% of patients with smoldering MM (SMM) and newly diagnosed MM (NDMM), and in around 70% of relapsed/refractory MM patients (RRMM).6 The impact of 1q21 on disease progression at an early stage has not been widely investigated. A few studies have suggested that the acquisition of extra 1q21 copies may play a role in disease progression.7,8 In fact, SMM patients with 1q21+ may be more likely to progress to MM than patients without 1q21+.8 Recent studies have demonstrated that the 1q21 copy number has a different impact on the responsiveness to MM treatments, especially proteasome inhibition (PI).9 PI is a well-established anti-cancer treatment approach used in MM. Throughout the years, the implementation of PI drugs as part of standard MM therapy has continued to improve the quality of life and clinical outcomes of MM patients. Furthermore, additional copies of 1q21 have been associated with PI resistance and recurrence of the disease in patients with 1q21+, limiting the long-term medical utility of PI.9,10 Recent studies have demonstrated that patients with 1q21+ treated with combination treatment with bortezomib (Bor) have inferior progression-free survival and overall survival compared to patients who do not present 1q21+.11 Similar results were observed when patients harboring 1q21 ampliSeveral genes are known to be deregulated upon the amplification of the 1q21 locus;9 nonetheless, the pathogenic and their possible role as druggable targets is not fully understood. In our study, we analyzed primary MM bone marrow (BM) PC from both SMM and NDMM patients to identify gene

Analysis of Gbx genes during neural development

[ACCESS RESTRICTED TO THE UNIVERSITY OF MISSOURI AT AUTHOR'S REQUEST.] The understanding of how transcription factors regulate neural patterning in the anterior hindbrain, and how the assembly of specific neuronal circuits controlling motor behavior in the developing spinal cord are important concerns in the field of developmental biology. In this dissertation we used two animal models, mouse and zebrafish, to study how gastrulation-brain homeobox (Gbx) class transcription factors (Gbx1 and Gbx2), affect the formation of sensorimotor circuits in mice and the development of anterior hindbrain in zebrafish. We use gene inactivation to study the role of Gbx1 and gbx2 during mouse and zebrafish embryogenesis. Gbx1 mutant (Gbx1-/-) mice develop a severe defect in motor coordination specifically affecting hindlimb gait. Loss of Gbx1 also resulted in an aberrant projection of the Ia proprioceptive sensory fiber in the ventral horn of the developing spinal cord. In addition, we show that Gbx1-/- embryos exhibit a reduction of Islet 1+ (ISL1) ventral motor neurons beginning at E14.5. In a parallel study, we investigated the role of gbx2 in the development of the anterior hindbrain in zebrafish embryos (Burroughs-Garcia et al., 2011). Gbx2 is widely known in mice and other vertebrates for it role positioning the mid hindbrain boundary. Despite many studies in mice showing the need of Gbx2 for normal hindbrain development, only few studies have focused on this role for Gbx2 in other vertebrates. Since the expression of Gbx2 in anterior hindbrain is conserved among vertebrate, we investigated whether the role of Gbx2 is functionally conserved during anterior hindbrain development in zebrafish. Loss-of-function studies using antisense morpholino show that gbx2 mutant embryos exhibit similar developmental defects as seen in hypomorphic mice (Waters and Lewandoski, 2006). We showed that deletion of gbx2 results in a truncation of the anterior hindbrain region between rhombomere (r)1 and r3 (Burroughs-Garcia et al., 2011). Furthermore there is an abnormal clustering of nV cell bodies arising from r2 and r3. These phenotypes can be rescued by injecting mouse Gbx2 mRNA (Buckley et al., 2013). This dissertation provides the first functional analysis of Gbx1 during mouse embryonic development. Additionally, our results provide evidence that gbx2/Gbx2 has shared an evolutionarily conserved role during the development of the anterior hindbrain in zebrafish embryos.Includes bibliographical references (pages 145-162)

Examination of Gbx2 function in zebrafish development [abstract]

Abstract only availableGbx2 is a member of the Gbx class of homeobox genes which encode DNA-binding transcription factors. The amino acid sequence of Gbx2 is highly conserved across multiple species (e.g. mice, zebrafish, chicken, and frogs) with 100% sequence identity between the Gbx2 homeodomains of the species examined. The function of Gbx2 in mice and zebrafish has been studied with loss-of-function and hypomorphic (reduced expression) models in both species. The results of these studies have demonstrated a requirement for Gbx2 in normal development of the mid/hindbrain organizer (isthmus) and anterior hindbrain. The hindbrain controls many basic life functions such as breathing and heartbeat. In early vertebrate development, the hindbrain is organized into eight distinct segments called rhombomeres. Rhombomeres give rise to hindbrain regions such as the cerebellum, pons, and medulla oblongata. In situ hybridization studies in Gbx2 null and hypomorphic mice have shown that cranial nerve V, which is derived from rhombomeres 2 and 3, fails to develop normally without wild-type levels of Gbx2. Mice lacking wild-type levels of Gbx2 die immediately after birth. To examine if Gbx2 has a similar impact on cranial nerve V development in zebrafish, we injected a morpholino specific for zebrafish Gbx2 into zebrafish embryos to silence the Gbx2 gene early in development. Our findings have shown that zebrafish subjected to the morpholino have a similar phenotype in the hindbrain as Gbx2 hypomorphic mice. Our present research will attempt to rescue the normal hindbrain phenotype in zebrafish embryos by simultaneously injecting the Gbx2 morpholino with synthesized zebrafish Gbx2 mRNA. We will also attempt to rescue the normal phenotype with synthesized mouse Gbx1 mRNA and mouse Gbx2 mRNA

The Metabolic Features of Osteoblasts: Implications for Multiple Myeloma (MM) Bone Disease

The study of osteoblast (OB) metabolism has recently received increased attention due to the considerable amount of energy used during the bone remodeling process. In addition to glucose, the main nutrient for the osteoblast lineages, recent data highlight the importance of amino acid and fatty acid metabolism in providing the fuel necessary for the proper functioning of OBs. Among the amino acids, it has been reported that OBs are largely dependent on glutamine (Gln) for their differentiation and activity. In this review, we describe the main metabolic pathways governing OBs' fate and functions, both in physiological and pathological malignant conditions. In particular, we focus on multiple myeloma (MM) bone disease, which is characterized by a severe imbalance in OB differentiation due to the presence of malignant plasma cells into the bone microenvironment. Here, we describe the most important metabolic alterations involved in the inhibition of OB formation and activity in MM patients

Characterization of the Gbx1-/- mouse mutant: a requirement for Gbx1 in normal locomotion and sensorimotor circuit development.

The Gbx class of homeobox genes encodes DNA binding transcription factors involved in regulation of embryonic central nervous system (CNS) development. Gbx1 is dynamically expressed within spinal neuron progenitor pools and becomes restricted to the dorsal mantle zone by embryonic day (E) 12.5. Here, we provide the first functional analysis of Gbx1. We generated mice containing a conditional Gbx1 allele in which exon 2 that contains the functional homeodomain is flanked with loxP sites (Gbx1(flox)); Cre-mediated recombination of this allele results in a Gbx1 null allele. In contrast to mice homozygous for a loss-of-function allele of Gbx2, mice homozygous for the Gbx1 null allele, Gbx1(-/-), are viable and reproductively competent. However, Gbx1(-/-) mice display a gross locomotive defect that specifically affects hindlimb gait. Analysis of embryos homozygous for the Gbx1 null allele reveals disrupted assembly of the proprioceptive sensorimotor circuit within the spinal cord, and a reduction in ISL1(+) ventral motor neurons. These data suggest a functional requirement for Gbx1 in normal development of the neural networks that contribute to locomotion. The generation of this null allele has enabled us to functionally characterize a novel role for Gbx1 in development of the spinal cord

Gbx1 and Gbx2 Are Essential for Normal Patterning and Development of Interneurons and Motor Neurons in the Embryonic Spinal Cord

The molecular mechanisms regulating neurogenesis involve the control of gene expression by transcription factors. Gbx1 and Gbx2, two members of the Gbx family of homeodomain-containing transcription factors, are known for their essential roles in central nervous system development. The expression domains of mouse Gbx1 and Gbx2 include regions of the forebrain, anterior hindbrain, and spinal cord. In the spinal cord, Gbx1 and Gbx2 are expressed in PAX2+ interneurons of the dorsal horn and ventral motor neuron progenitors. Based on their shared domains of expression and instances of overlap, we investigated the functional relationship between Gbx family members in the developing spinal cord using Gbx1&minus;/&minus;, Gbx2&minus;/&minus;, and Gbx1&minus;/&minus;/Gbx2&minus;/&minus; embryos. In situ hybridization analyses of embryonic spinal cords show upregulation of Gbx2 expression in Gbx1&minus;/&minus; embryos and upregulation of Gbx1 expression in Gbx2&minus;/&minus; embryos. Additionally, our data demonstrate that Gbx genes regulate development of a subset of PAX2+ dorsal inhibitory interneurons. While we observe no difference in overall proliferative status of the developing ependymal layer, expansion of proliferative cells into the anatomically defined mantle zone occurs in Gbx mutants. Lastly, our data shows a marked increase in apoptotic cell death in the ventral spinal cord of Gbx mutants during mid-embryonic stages. While our studies reveal that both members of the Gbx gene family are involved in development of subsets of PAX2+ dorsal interneurons and survival of ventral motor neurons, Gbx1 and Gbx2 are not sufficient to genetically compensate for the loss of one another. Thus, our studies provide novel insight to the relationship harbored between Gbx1 and Gbx2 in spinal cord development