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Full Abstracts – GAMETOGENESIS AND ORGANOGENESIS
173
149
Muscle size and myonuclear position are independently regulated by distinct Dynein pathways.
Victoria K. Schulman
1,2
, Eric S. Folker
2
, Mary K.
Baylies
1,2
. 1) Weill Cornell Graduate School of Medical Sciences, New York, NY 10065; 2) Sloan-Kettering Institute, Memorial Sloan-Kettering Cancer
Center, New York, NY 10065.
Morphogenesis, through modulation of the cytoskeleton, dictates cell size, shape, and organization, and informs cellular function. Patients with muscle
diseases present with aberrant muscle cell morphologies characterized by smaller myofibers with mispositioned nuclei. However, the mechanisms that
control these processes and their contribution to muscle weakness in disease are not known. To understand how the cytoskeleton impacts muscle form and
function, we examined the role of Dynein during
Drosophila
muscle development and found that regulation of muscle size and myonuclear positioning are
mechanistically distinct. Several factors including the Dynein heavy chain (Dhc64C), the Dynein light chain (Dlc90F), and Partner of Inscuteable (Pins)
contribute to both muscle growth and myonuclear positioning. However, Lis1 contributes only to Dynein-dependent muscle size, whereas CLIP-190 and
Glued only contribute to Dynein-dependent myonuclear positioning. Moreover, Lis1 and CLIP-190 do not functionally interact. Mechanistically, there is a
decrease in the density of microtubules at the muscle poles in
clip190
mutants, suggesting that microtubule interactions with the cortex are required for
proper nuclear positioning. In
lis1
mutants, Dynein hyper-accumulates at the muscle poles, suggesting that retrograde trafficking away from the poles is
required for proper muscle growth. Importantly, all mutants had the correct number of nuclei present within each myofiber, suggesting that fusion alone is
not sufficient to regulate myofiber size. The effects of both Lis1 and CLIP-190 are downstream of Dynein arriving at the muscle pole, suggesting that these
proteins specify separate Dynein functions within a single localization. Finally, defects in muscle size or myonuclear positioning impair muscle function
in
vivo
. These findings indicate that muscle size and myonuclear positioning are essential for muscle function, yet regulated by distinct Dynein-dependent
mechanisms.
150
MiR-92b is a heart and muscle specific microRNA that regulates Mef2 level through a negative feedback loop in
Drosophila
.
Zhimin Chen
1
,
Shanshan Liang
1
, Ying Zhao
1
, Zhe Han
1,2
. 1) Department of Internal Medicine, Division of Molecular Medicine and Genetics, University of Michigan
Medical School, Ann Arbor, MI; 2) Department of Cell and Developmental Biology, University of Michigan Medical School, Ann Arbor, MI.
Level of Mef2 transcription factor must be precisely controlled since Mef2 is essential for heart and muscle differentiation by activating hundreds of target
genes. Since Mef2 can self-activate, a negative feedback loop must present to keep its expression level stable. Using microarray and genetic analysis, we
have identified miR-92b, a highly conserved microRNA, to be a post-transcriptional regulator of Mef2 in
Drosophila
. MiR-92b is expressed in a heart and
muscle specific pattern similar to that of Mef2 and is directly activated by Mef2 through conserved Mef2 binding sites in its
cis
-regulatory region. On the
other hand, miR-92b represses Mef2 translation through binding to conserved miR-92 binding sites in the 3’ UTR of Mef2, forming a negative feedback
loop that maintains Mef2 level in a proper range. Perturbing this negative feedback loop, either by deleting or over-expressing miR-92b, causes
corresponding up or down changes of Mef2 protein level and affects muscle development. A microRNA usually has many targets and its level needs to be
tightly controlled as well. Our data suggest that a microRNA and a transcription factor can form a negative feedback loop to maintain the stable expression
level of both efficiently.
151
Mitotic Cell Rounding Accelerates Invagination of the
Drosophila
Tracheal Placode.
Takefumi Kondo, Shigeo Hayashi. Lab. for morphogenetic
signaling, RIKEN CDB, Kobe, Hyogo, Japan.
Animal cells change their shape to sphere upon entry into mitotic phase. Mitotic cell rounding is a conserved process governed by extensive rearrangement
of actin cytoskeleton, and is necessary for proper cell division. In addition, mitosis is also coupled with microtubule reorganization to form spindle. During
development, since the regulation of cytoskeletal structure in interphase is involved in cell shape change and modulation of cell-cell adhesion, mitotic entry
must be precisely controlled to avoid interference of tissue morphogenesis and integrity. On the other hand, whether mitotic cell rounding itself plays an
active role in epithelial morphogenesis is unknown.
Invagination is one of the key morphogenetic processes, which converts flat epithelial sheets into three-dimensional structures. To understand cellular
mechanisms that accomplish the full process of epithelial invagination, we performed three-dimensional live imaging of
Drosophila
tracheal placode as a
model system, and found that this morphogenetic event is divided into two distinct phases by speed. First, the slow invagination phase proceeds with a wave
of circular Myosin concentration formed in the placode, causing a series of cell intercalation and apical constriction of central cells. In the second phase,
speed of invagination was suddenly increased. This acceleration was associated with entry of one of the central cells into mitosis and rounding at the basal
side, resulting in the fast basal movement of the apical surface of the placode. Genetic block of mitotic entry and cell rounding interrupted transition to the
second phase. On the other hand, microtubule inhibitor colchicine, which prevents spindle formation and cell division, but not cell rounding, did not interfere
the phase transition. These results indicate that cell rounding accompanied by mitotic entry actively drives fast invagination of tracheal placode.