TyAnna L. Lovato
University of New Mexico
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Featured researches published by TyAnna L. Lovato.
Developmental Biology | 2012
Anton L. Bryantsev; Phillip W. Baker; TyAnna L. Lovato; MaryAnn S. Jaramillo; Richard M. Cripps
Identifying the genetic program that leads to formation of functionally and morphologically distinct muscle fibers is one of the major challenges in developmental biology. In Drosophila, the Myocyte Enhancer Factor-2 (MEF2) transcription factor is important for all types of embryonic muscle differentiation. In this study we investigated the role of MEF2 at different stages of adult skeletal muscle formation, where a diverse group of specialized muscles arises. Through stage- and tissue-specific expression of Mef2 RNAi constructs, we demonstrate that MEF2 is critical at the early stages of adult myoblast fusion: mutant myoblasts are attracted normally to their founder cell targets, but are unable to fuse to form myotubes. Interestingly, ablation of Mef2 expression at later stages of development showed MEF2 to be more dispensable for structural gene expression: after myoblast fusion, Mef2 knockdown did not interrupt expression of major structural gene transcripts, and myofibrils were formed. However, the MEF2-depleted fibers showed impaired integrity and a lack of fibrillar organization. When Mef2 RNAi was induced in muscles following eclosion, we found no adverse effects of attenuating Mef2 function. We conclude that in the context of adult myogenesis, MEF2 remains an essential factor, participating in control of myoblast fusion, and myofibrillogenesis in developing myotubes. However, MEF2 does not show a major requirement in the maintenance of muscle structural gene expression. Our findings point to the importance of a diversity of regulatory factors that are required for the formation and function of the distinct muscle fibers found in animals.
Insect Molecular Biology | 2001
TyAnna L. Lovato; S. M. Meadows; Phillip W. Baker; J. C. Sparrow; Richard M. Cripps
Actin is a ubiquitous and highly conserved eukaryotic protein required for cell motility and locomotion. In this manuscript, we characterize the four muscle actin genes of the insect Drosophila virilis and demonstrate strong similarities between the D. virilis genes and their homologues in Drosophila melanogaster; intron locations are conserved, and there are few amino acid differences between homologues. We also found strong conservation in temporal expression patterns of the muscle actin genes – the homologues of the D. melanogaster genes Act57B and Act87E are expressed throughout the life cycle, whereas the other two D. virilis genes, homologous to Act79B and Act88F are specific to pupal and adult stages. In situ hybridization revealed that each D. virilis gene is expressed in a unique pattern in the muscles of the thorax and abdomen. These muscle‐specific patterns of actin isoforms suggest a greater physiological diversity for the adult muscles of insects than has been appreciated to date from their categorization into fibrillar, tubular (non‐fibrillar) and supercontractile muscle types.
Developmental Cell | 2012
Anton L. Bryantsev; Sandy Duong; Tonya M. Brunetti; Maria B. Chechenova; TyAnna L. Lovato; Cloyce Nelson; Elizabeth Shaw; Juli D. Uhl; Brian Gebelein; Richard M. Cripps
Here we identify a key role for the homeodomain proteins Extradenticle (Exd) and Homothorax (Hth) in the specification of muscle fiber fate in Drosophila. exd and hth are expressed in the fibrillar indirect flight muscles but not in tubular jump muscles, and manipulating exd or hth expression converts one muscle type into the other. In the flight muscles, exd and hth are genetically upstream of another muscle identity gene, salm, and are direct transcriptional regulators of the signature flight muscle structural gene, Actin88F. Exd and Hth also impact muscle identity in other somatic muscles of the body by cooperating with Hox factors. Because mammalian orthologs of exd and hth also contribute to muscle gene regulation, our studies suggest that an evolutionarily conserved genetic pathway determines muscle fiber differentiation.
Methods of Molecular Biology | 2012
Ginny R. Morriss; Anton L. Bryantsev; Maria B. Chechenova; Elisa M. LaBeau; TyAnna L. Lovato; Kathryn M. Ryan; Richard M. Cripps
The Drosophila system has been invaluable in providing important insights into mesoderm specification, muscle specification, myoblast fusion, muscle differentiation, and myofibril assembly. Here, we present a series of Drosophila protocols that enable the researcher to visualize muscle precursors and differentiated muscles, at all stages of development. In doing so, we also highlight the variety of techniques that are used to create these findings. These protocols are directly used for the Drosophila system, and are provided with explanatory detail to enable the researcher to apply them to other systems.
Biochemistry and Molecular Biology Education | 2016
Vanesa Adame; Holly Chapapas; Marilyn Cisneros; Carol Deaton; Sophia Deichmann; Chauncey Gadek; TyAnna L. Lovato; Maria B. Chechenova; Paul Guerin; Richard M. Cripps
CRISPR/Cas9 genome editing technology is used in the manipulation of genome sequences and gene expression. Because of the ease and rapidity with which genes can be mutated using CRISPR/Cas9, we sought to determine if a single‐semester undergraduate class could be successfully taught, wherein students isolate mutants for specific genes using CRISPR/Cas9. Six students were each assigned a single Drosophila gene, for which no mutants currently exist. Each student designed and created plasmids to encode single guide RNAs that target their selected gene; injected the plasmids into Cas9‐expressing embryos, in order to delete the selected gene; carried out a three‐generation cross to test for germline transmission of a mutated allele and generate a stable stock of the mutant; and characterized the mutant alleles by PCR and sequencing. Three genes out of six were successfully mutated. Pre‐ and post‐ survey evaluations of the students in the class revealed that student attitudes towards their research competencies increased, although the changes were not statistically significant. We conclude that it is feasible to develop a laboratory genome editing class, to provide effective laboratory training to undergraduate students, and to generate mutant lines for use by the broader scientific community.
Genetics | 2009
TyAnna L. Lovato; Melanie M. Adams; Phillip W. Baker; Richard M. Cripps
Temperature-sensitive (TS) mutations are a useful tool for elucidating gene function where a gene of interest is essential at multiple stages of development. However, the molecular mechanisms behind TS alleles vary. TS mutations of the myogenic regulator Myocyte enhancer factor-2 (MEF2) in Drosophila arise in the heteroallelic combination Mef230-5/Mef244-5. We show that the 30-5 mutation affects the N-terminal MADS domain, causing impaired DNA binding ability and failure of homozygous mutants to survive to adulthood. The 44-5 mutation deletes a downstream splice acceptor site and results in a truncated protein that is unable to activate MEF2 targets. 44-5 homozygotes consequently show severely impaired myogenesis and die as embryos. We propose that in heteroallelic mutants at the permissive temperature, 30-5/44-5 heterodimers form and have a sufficiently stable interaction with DNA to activate myogenic gene expression; at the restrictive temperature, 44-5 homodimers displace 30-5/44-5 heterodimers from target genes, thus acting in a dominant-negative manner. To test this, we show that 30-5/44-5 heterodimers can form, and we study additional Mef2 alleles for complementation with the 30-5 allele. An allele affecting the DNA binding domain fails to complement 30-5, whereas two alleles affecting downstream residues show temperature-dependent complementation. Thus, by combining one MEF2 isoform having weakened DNA binding ability with a second truncated MEF2 mutant that has lost its activation ability, a TS form of intragenic complementation can be generated. These findings will provide new insight and guidance into the functions of dimeric proteins and how they might be engineered to generate TS combinations.
Developmental Biology | 2016
Gloriana V. Trujillo; Dalea H. Nodal; Candice V. Lovato; Jill Hendren; Lynda A. Helander; TyAnna L. Lovato; Rolf Bodmer; Richard M. Cripps
The inflow tracts of the embryonic Drosophila cardiac tube, termed ostia, arise in its posterior three segments from cardiac cells that co-express the homeotic transcription factor Abdominal-A (abdA), the orphan nuclear receptor Seven-up (Svp), and the signaling molecule Wingless (Wg). To define the roles of these factors in inflow tract development, we assessed their function in inflow tract formation. We demonstrate, using several criteria, that abdA, svp, and wg are each critical for normal inflow tract formation. We further show that Wg acts in an autocrine manner to impact ostia fate, and that it mediates this effect at least partially through the canonical Wg signaling pathway. By contrast, neither wg expression nor Wg signaling are sufficient for inflow tract formation when expressed in anterior Svp cells that do not normally form inflow tracts in the embryo. Instead, ectopic abd-A expression throughout the cardiac tube is required for the formation of ectopic inflow tracts, indicating that autocrine Wg signaling must be supplemented by additional Hox-dependent factors to effect inflow tract formation. Taken together, these studies define important cellular and molecular events that contribute to cardiac inflow tract development in Drosophila. Given the broad conservation of the cardiac regulatory network through evolution, our studies provide insight into mechanisms of cardiac development in higher animals.
PLOS ONE | 2015
TyAnna L. Lovato; Cheryl A. Sensibaugh; Kirstie L. Swingle; Melody M. Martinez; Richard M. Cripps
Expression of the MADS domain transcription factor Myocyte Enhancer Factor 2 (MEF2) is regulated by numerous and overlapping enhancers which tightly control its transcription in the mesoderm. To understand how Mef2 expression is controlled in the heart, we identified a late stage Mef2 cardiac enhancer that is active in all heart cells beginning at stage 14 of embryonic development. This enhancer is regulated by the NK-homeodomain transcription factor Tinman, and the GATA transcription factor Pannier through both direct and indirect interactions with the enhancer. Since Tinman, Pannier and MEF2 are evolutionarily conserved from Drosophila to vertebrates, and since their vertebrate homologs can convert mouse fibroblast cells to cardiomyocytes in different activator cocktails, we tested whether over-expression of these three factors in vivo could ectopically activate known cardiac marker genes. We found that mesodermal over-expression of Tinman and Pannier resulted in approximately 20% of embryos with ectopic Hand and Sulphonylurea receptor (Sur) expression. By adding MEF2 alongside Tinman and Pannier, a dramatic expansion in the expression of Hand and Sur was observed in almost all embryos analyzed. Two additional cardiac markers were also expanded in their expression. Our results demonstrate the ability to initiate ectopic cardiac fate in vivo by the combination of only three members of the conserved Drosophila cardiac transcription network, and provide an opportunity for this genetic model system to be used to dissect the mechanisms of cardiac specification.
Archive | 2019
Anton L. Bryantsev; Lizzet Castillo; Sandy T. Oas; Maria B. Chechenova; Tracy E. Dohn; TyAnna L. Lovato
Drosophila is a useful model organism for studying the molecular signatures that define specific muscle types during myogenesis. It possesses significant genetic conservation with humans for muscle disease causing genes and a lack of redundancy that simplifies functional analysis. Traditional molecular methods can be utilized to understand muscle developmental processes such as Western blots, in situ hybridizations, RT-PCR and RNAseq, to name a few. However, one challenge for these molecular methods is the ability to dissect different muscle types. In this protocol we describe some useful techniques for extracting muscles from the pupal and adult stages of development using flight and jump muscles as an example.
Circulation-cardiovascular Genetics | 2017
TyAnna L. Lovato; Richard M. Cripps
In 1925, Dr Carlos Monge presented to the National Academy of Medicine in Lima, Peru, his observations of a new disease. His patient had bluish skin, dizziness, and confusion when working at high altitudes, but symptoms were ameliorated once the patient returned to the coast.1 In a storied and impactful career, Monge went on to study the physiological differences between sufferers of Monge’s disease, those who were able to acclimatize to the altitude, and finally those who thrived in the low oxygen conditions.2 Today, we now know this condition as chronic mountain sickness (CMS). Caused by exposure to low oxygen conditions, CMS affects a significant proportion of high-altitude populations and can lead to pulmonary hypertension, cardiac hypertrophy, and eventual heart failure.3 See Article by Zarndt and Walls et al Today, CMS still afflicts a large number of individuals, including a striking one sixth of residents of Cerro de Pasco in Peru,4 designating CMS as a significant medical challenge in many high-altitude populations. Understanding the mechanisms by which hypertension and cardiac hypertrophy occur as a result of hypoxia is important not only …