Andrew Muroyama

Bio: Andrew Muroyama is an academic researcher from Duke University. The author has contributed to research in topics: Microtubule & Centrosome. The author has an hindex of 7, co-authored 12 publications receiving 418 citations. Previous affiliations of Andrew Muroyama include Stanford University.

Microtubule organization, dynamics and functions in differentiated cells.

Abstract: Over the past several decades, numerous studies have greatly expanded our knowledge about how microtubule organization and dynamics are controlled in cultured cells in vitro However, our understanding of microtubule dynamics and functions in vivo, in differentiated cells and tissues, remains under-explored. Recent advances in generating genetic tools and imaging technologies to probe microtubules in situ, coupled with an increased interest in the functions of this cytoskeletal network in differentiated cells, are resulting in a renaissance. Here, we discuss the lessons learned from such approaches, which have revealed that, although some differentiated cells utilize conserved strategies to remodel microtubules, there is considerable diversity in the underlying molecular mechanisms of microtubule reorganization. This highlights a continued need to explore how differentiated cells regulate microtubule geometry in vivo.

NuMA-microtubule interactions are critical for spindle orientation and the morphogenesis of diverse epidermal structures

Abstract: Before a cell divides, it must duplicate its DNA so that each new cell receives a complete set of genetic material. A structure called the mitotic spindle helps to ensure each new cell gets the correct amount of DNA. Cells often precisely position their mitotic spindle during division, and this spindle orientation is important for generating different types of cells and for establishing the three-dimensional structure of tissues. How cells rotate their spindles into the correct position is not well understood, but a protein called NuMA is important for this process. Seldin et al. developed genetic tools that could disrupt spindle orientation in specific types of cells to determine where this orientation is important for proper tissue development. This revealed that the correct placement of the mitotic spindle is important for the development of the skin of mouse embryos and the formation of the hair of adult mice. Seldin et al. also found that the NuMA protein binds to the tips of the microtubules that make up the mitotic spindle. This binding activity is important for NuMA to be able to position the mitotic spindle correctly in the cell. The findings suggest similarities between how cells orient mitotic spindles and how they segregate DNA during cell division. More work is now needed to better understand how NuMA collaborates with force-generating molecular motors to precisely orient the mitotic spindle in the cell. In addition, understanding how spindle orientation dictates the fate of cells in the skin is an important future goal.

Divergent regulation of functionally distinct γ-tubulin complexes during differentiation

Abstract: Differentiation induces the formation of noncentrosomal microtubule arrays in diverse tissues. The formation of these arrays requires loss of microtubule-organizing activity (MTOC) at the centrosome, but the mechanisms regulating this transition remain largely unexplored. Here, we use the robust loss of centrosomal MTOC activity in the epidermis to identify two pools of γ-tubulin that are biochemically and functionally distinct and differentially regulated. Nucleation-competent CDK5RAP2-γ-tubulin complexes were maintained at centrosomes upon initial epidermal differentiation. In contrast, Nedd1-γ-tubulin complexes did not promote nucleation but were required for anchoring of microtubules, a previously uncharacterized activity for this complex. Cell cycle exit specifically triggered loss of Nedd1-γ-tubulin complexes, providing a mechanistic link connecting MTOC activity and differentiation. Collectively, our studies demonstrate that distinct γ-tubulin complexes regulate different microtubule behaviors at the centrosome and show that differential regulation of these complexes drives loss of centrosomal MTOC activity.

Actin-related protein2/3 complex regulates tight junctions and terminal differentiation to promote epidermal barrier formation

Abstract: The epidermis provides an essential seal from the external environment and retains fluids within the body. To form an effective barrier, cells in the epidermis must form tight junctions and terminally differentiate into cornified envelopes. Here, we demonstrate that the branched actin nucleator, the actin-related protein (Arp)2/3 complex, is unexpectedly required for both these activities. Loss of the ArpC3 subunit of the Arp2/3 complex resulted in minimal changes in the morphogenesis and architecture of this stratified squamous epithelium, but resulted in profound defects in its physiology. Mutant embryos did not develop an effective barrier to the external environment and died within hours of birth. We discovered two underlying causes for these effects. First, ArpC3 was essential for robust assembly and function of tight junctions, specialized cell–cell adhesions that restrict water loss in the epidermis. Second, there were defects in differentiation of the epidermis and the production of cornified envelopes, structures essential for barrier activity. Underlying this defect, we found that YAP was inappropriately active not only in the ArpC3 mutant tissue, but also in cultured cells. Inhibition of YAP activity rescued the differentiation and barrier defects caused by loss of ArpC3. These results demonstrate previously unappreciated roles for the Arp2/3 complex and highlight the functions of branched actin networks in a complex tissue.

A non-apoptotic cell death process, entosis, that occurs by cell-in-cell invasion.

Abstract: 1560 Epithelial cells require attachment to extra-cellular matrix (ECM) to suppress an apoptotic cell death program termed anoikis. We describe a non-apoptotic cell death program in matrix-detached cells, termed entosis, that is initiated by a previously unrecognized and unusual process involving the invasion of one cell into another, leading to a transient state in which a live cell is contained within a neighboring host cell. These 9cell-in-cell9 structures closely resemble similar cytological features in human cancers. Although a small percentage of live internalized cells can be released, the majority of internalized cells undergo non-apoptotic cell death by lysosomal acidification. Based on these data, we present evidence for a tumor suppressive role of entosis in human breast cancer. Paradoxically, we have also found that entosis can promote the development of aneuploidy, as live internalized cells can disrupt cytokinesis of host cells leading to multinucleation in vitro and in vivo. Thus, entosis may be tumor suppressive in some contexts and paradoxically tumor promoting in others. Models to investigate the role of entosis in human tumors will be presented.

Regulation of mitotic spindle orientation: an integrated view.

Abstract: Mitotic spindle orientation is essential for cell fate decisions, epithelial maintenance, and tissue morphogenesis. In most animal cell types, the dynein motor complex is anchored at the cell cortex and exerts pulling forces on astral microtubules to position the spindle. Early studies identified the evolutionarily conserved Gαi/LGN/NuMA complex as a key regulator that polarizes cortical force generators. In recent years, a combination of genetics, biochemistry, modeling, and live imaging has contributed to decipher the mechanisms of spindle orientation. Here, we highlight the dynamic nature of the assembly of this complex and discuss the molecular regulation of its localization. Remarkably, a number of LGN-independent mechanisms were described recently, whereas NuMA remains central in most pathways involved in recruiting force generators at the cell cortex. We also describe the emerging role of the actin cortex in spindle orientation and discuss how dynamic astral microtubule formation is involved. We further give an overview on instructive external signals that control spindle orientation in tissues. Finally, we discuss the influence of cell geometry and mechanical forces on spindle orientation.

Coiled‐coils: The long and short of it

Abstract: Coiled‐coils are found in proteins throughout all three kingdoms of life. Coiled‐coil domains of some proteins are almost invariant in sequence and length, betraying a structural and functional role for amino acids along the entire length of the coiled‐coil. Other coiled‐coils are divergent in sequence, but conserved in length, thereby functioning as molecular spacers. In this capacity, coiled‐coil proteins influence the architecture of organelles such as centrioles and the Golgi, as well as permit the tethering of transport vesicles. Specialized coiled‐coils, such as those found in motor proteins, are capable of propagating conformational changes along their length that regulate cargo binding and motor processivity. Coiled‐coil domains have also been identified in enzymes, where they function as molecular rulers, positioning catalytic activities at fixed distances. Finally, while coiled‐coils have been extensively discussed for their potential to nucleate and scaffold large macromolecular complexes, structural evidence to substantiate this claim is relatively scarce.