<html xmlns:v="urn:schemas-microsoft-com:vml" xmlns:o="urn:schemas-microsoft-com:office:office" xmlns:w="urn:schemas-microsoft-com:office:word" xmlns:m="http://schemas.microsoft.com/office/2004/12/omml" xmlns="http://www.w3.org/TR/REC-html40"><head><meta http-equiv=Content-Type content="text/html; charset=utf-8"><meta name=Generator content="Microsoft Word 14 (filtered medium)"><style><!--
/* Font Definitions */
@font-face
{font-family:"Cambria Math";
panose-1:2 4 5 3 5 4 6 3 2 4;}
@font-face
{font-family:Cambria;
panose-1:2 4 5 3 5 4 6 3 2 4;}
@font-face
{font-family:Calibri;
panose-1:2 15 5 2 2 2 4 3 2 4;}
@font-face
{font-family:Tahoma;
panose-1:2 11 6 4 3 5 4 4 2 4;}
@font-face
{font-family:"Franklin Gothic Book";
panose-1:2 11 5 3 2 1 2 2 2 4;}
/* Style Definitions */
p.MsoNormal, li.MsoNormal, div.MsoNormal
{margin:0in;
margin-bottom:.0001pt;
font-size:11.0pt;
font-family:"Calibri","sans-serif";}
a:link, span.MsoHyperlink
{mso-style-priority:99;
color:blue;
text-decoration:underline;}
a:visited, span.MsoHyperlinkFollowed
{mso-style-priority:99;
color:purple;
text-decoration:underline;}
p.MsoAcetate, li.MsoAcetate, div.MsoAcetate
{mso-style-priority:99;
mso-style-link:"Balloon Text Char";
margin:0in;
margin-bottom:.0001pt;
font-size:8.0pt;
font-family:"Tahoma","sans-serif";}
span.BalloonTextChar
{mso-style-name:"Balloon Text Char";
mso-style-priority:99;
mso-style-link:"Balloon Text";
font-family:"Tahoma","sans-serif";}
span.EmailStyle19
{mso-style-type:personal;
font-family:"Calibri","sans-serif";
color:windowtext;}
span.EmailStyle20
{mso-style-type:personal;
font-family:"Calibri","sans-serif";
color:#1F497D;}
span.EmailStyle21
{mso-style-type:personal;
font-family:"Calibri","sans-serif";
color:#1F497D;}
span.EmailStyle22
{mso-style-type:personal;
font-family:"Calibri","sans-serif";
color:#1F497D;}
span.EmailStyle23
{mso-style-type:personal;
font-family:"Calibri","sans-serif";
color:#1F497D;}
span.EmailStyle24
{mso-style-type:personal;
font-family:"Calibri","sans-serif";
color:#1F497D;}
span.EmailStyle25
{mso-style-type:personal;
font-family:"Calibri","sans-serif";
color:#1F497D;}
span.EmailStyle26
{mso-style-type:personal;
font-family:"Calibri","sans-serif";
color:#1F497D;}
span.EmailStyle27
{mso-style-type:personal;
font-family:"Calibri","sans-serif";
color:#1F497D;}
span.EmailStyle28
{mso-style-type:personal;
font-family:"Calibri","sans-serif";
color:#1F497D;}
span.EmailStyle29
{mso-style-type:personal;
font-family:"Calibri","sans-serif";
color:#1F497D;}
span.EmailStyle30
{mso-style-type:personal;
font-family:"Calibri","sans-serif";
color:#1F497D;}
span.EmailStyle31
{mso-style-type:personal;
font-family:"Calibri","sans-serif";
color:#1F497D;}
span.EmailStyle32
{mso-style-type:personal;
font-family:"Calibri","sans-serif";
color:#1F497D;}
span.EmailStyle33
{mso-style-type:personal;
font-family:"Calibri","sans-serif";
color:#1F497D;}
span.EmailStyle34
{mso-style-type:personal;
font-family:"Calibri","sans-serif";
color:#1F497D;}
span.EmailStyle35
{mso-style-type:personal;
font-family:"Calibri","sans-serif";
color:#1F497D;}
span.EmailStyle36
{mso-style-type:personal;
font-family:"Calibri","sans-serif";
color:#1F497D;}
span.EmailStyle37
{mso-style-type:personal;
font-family:"Calibri","sans-serif";
color:#1F497D;}
span.EmailStyle38
{mso-style-type:personal;
font-family:"Calibri","sans-serif";
color:#1F497D;}
span.EmailStyle39
{mso-style-type:personal;
font-family:"Calibri","sans-serif";
color:#1F497D;}
span.EmailStyle40
{mso-style-type:personal;
font-family:"Calibri","sans-serif";
color:#1F497D;}
span.EmailStyle41
{mso-style-type:personal;
font-family:"Calibri","sans-serif";
color:#1F497D;}
span.EmailStyle42
{mso-style-type:personal;
font-family:"Calibri","sans-serif";
color:#1F497D;}
span.EmailStyle43
{mso-style-type:personal-reply;
font-family:"Calibri","sans-serif";
color:#1F497D;}
span.apple-style-span
{mso-style-name:apple-style-span;}
.MsoChpDefault
{mso-style-type:export-only;
font-size:10.0pt;}
@page WordSection1
{size:8.5in 11.0in;
margin:1.0in 1.0in 1.0in 1.0in;}
div.WordSection1
{page:WordSection1;}
--></style><!--[if gte mso 9]><xml>
<o:shapedefaults v:ext="edit" spidmax="1026" />
</xml><![endif]--><!--[if gte mso 9]><xml>
<o:shapelayout v:ext="edit">
<o:idmap v:ext="edit" data="1" />
</o:shapelayout></xml><![endif]--></head><body lang=EN-US link=blue vlink=purple><div class=WordSection1><p class=MsoNormal><o:p> </o:p></p><p class=MsoNormal><o:p> </o:p></p><p class=MsoNormal><span style='font-size:14.0pt'><o:p> </o:p></span></p><p class=MsoNormal align=center style='margin-bottom:12.0pt;text-align:center'><b><span style='font-size:14.0pt'>UH ChBE Dept. Seminar<br>9:00am-10:00am, Tuesday , February 16, 2016<br>Cemo Hall, room 105</span></b><span style='font-size:14.0pt'><o:p></o:p></span></p><p class=MsoNormal align=center style='text-align:center'><span style='font-size:20.0pt;font-family:"Times New Roman","serif"'>Developing Novel Single-Cell Sequencing Technologies to Study Cellular Heterogeneity<o:p></o:p></span></p><p class=MsoNormal align=center style='text-align:center'><b><span style='font-size:14.0pt'>Siddharth Dey<o:p></o:p></span></b></p><p class=MsoNormal align=center style='text-align:center'><b><span style='font-size:14.0pt'>Hubrecht Institute, The Netherlands<o:p></o:p></span></b></p><p class=MsoNormal align=center style='text-align:center'><b><o:p> </o:p></b></p><p class=MsoNormal><b><span style='font-size:14.0pt'><o:p> </o:p></span></b></p><p class=MsoNormal><o:p> </o:p></p><p class=MsoNormal style='text-align:justify'><b><u><span style='font-size:14.0pt'>ABSTRACT:</span></u></b><span style='font-size:14.0pt'> </span><span style='font-size:10.5pt;font-family:"Arial","sans-serif"'>A</span><span style='font-size:10.5pt;font-family:"Arial","sans-serif"'> central question in biology is to understand how variability in the genome or epigenome regulates gene expression heterogeneity, thereby influencing cellular functions. </span><span style='font-size:10.5pt;font-family:"Arial","sans-serif"'>While heterogeneity in gene expression between individual cells has been studied extensively, </span><span style='font-size:10.5pt;font-family:"Arial","sans-serif"'>with important biological consequences ranging from drug resistance in bacteria and tumors to biasing lineage choices in differentiating mammalian progenitor cells,</span><span style='font-size:10.5pt;font-family:"Arial","sans-serif"'> the upstream mechanisms regulating cell-to-cell heterogeneity have been difficult to study and are poorly understood. To elucidate this relationship, I will be describing three studies where we developed novel single-cell technologies to understand how the genome and epigenome regulates the dynamics of gene expression. To understand how the local chromatin environment regulates gene expression variability, we </span><span style='font-size:10.5pt;font-family:"Arial","sans-serif"'>combined stochastic modeling and DNA accessibility measurements with single-molecule mRNA FISH to show that more repressed chromatin is associated with increased heterogeneity in gene expression. These results have important implications for understanding viral latency. Next, we <span style='letter-spacing:-.05pt'>developed the first genome-wide technology that enabled sequencing both genomic DNA and mRNA from the same cell. We used this technology to show that low genomic copy number regions are associated with increased gene expression variability between individual tumor cells, suggesting that copy number variations may be a mechanism to tune cell-to-cell heterogeneity in gene expression. Finally, I will describe a new genome-wide single-cell method for quantifying the DNA modification 5-hydroxymethylcytosine (5hmC), which has recently been shown to have critical roles in development and gene regulation. Using this method, we discovered pronounced cell-to-cell variability </span>in the relative amounts of 5hmC on the two DNA strands of a chromosome in mouse embryonic stem cells and 2-cell mouse embryos. By combining these results with a stochastic model, we demonstrated that even isogenic cell populations exposed to identical environments can display pronounced chromosome-wide epigenetic heterogeneity due to the slow kinetics of 5hmC turnover. Thus, these studies are beginning to reveal fundamental new insights into how variability in the genome or epigenome <span class=apple-style-span><span style='background:white'>influences cellular heterogeneity and ultimately, cellular phenotypes. Finally, I will briefly discuss future research plans where I propose to develop</span></span> </span><span style='font-size:10.5pt;font-family:"Arial","sans-serif";color:#222222;background:white'>novel integrated single-cell sequencing technologies that enable simultaneous genome-wide measurements of the epigenome and transcriptome from the same cell to gain insights into tumor evolution and progression, early mammalian development, and mechanisms regulating the maintenance and regeneration of adult tissues, knowledge that will be critical to more efficiently direct differentiation for regenerative medicine applications. </span><span style='font-size:10.5pt;font-family:"Arial","sans-serif"'><o:p></o:p></span></p><p class=MsoNormal style='text-align:justify'><span style='font-size:14.0pt;font-family:"Times New Roman","serif";color:black'><o:p> </o:p></span></p><p class=MsoNormal style='text-align:justify;background:white'><span style='font-family:"Cambria","serif";color:#1F497D'><o:p> </o:p></span></p><p class=MsoNormal style='margin-bottom:8.0pt;text-align:justify;text-indent:.5in'><b><u><span style='font-size:14.0pt'>BIO: </span></u></b><span style='font-size:10.5pt;font-family:"Arial","sans-serif"'>Siddharth Dey graduated from the Institute of Chemical Technology (Formerly: UDCT), Mumbai in 2006 with a B.S. in Chemical Engineering. Next, he worked with Professor David Schaffer and Professor Adam Arkin and received his Ph.D. in Chemical and Biomolecular Engineering in 2012 from the University of California, Berkeley. He has been interested in understanding how genetic and epigenetic factors regulate cell-to-cell variability in gene expression and influence cellular phenotypes. Specifically, during his graduate studies, he used quantitative single-cell techniques to investigate how the decision between a latent and lytic state in Human Immunodeficiency Virus-1 (HIV-1) is regulated by gene expression variability. In 2012, he moved to Professor Alexander van Oudenaarden's group as a post-doctoral researcher at the Hubrecht Institute, The Netherlands. Expanding upon this graduate work to study how upstream regulatory factors influence gene expression variability on a genome-wide scale, he started developing novel single-cell genomics technologies. He developed the first integrated single-cell method that enabled sequencing both genomic DNA and mRNA from the same cell. Further, he has been working on developing new methods to quantify epigenetic marks such as DNA methylation and </span><span style='font-size:10.5pt;font-family:"Arial","sans-serif";letter-spacing:-.05pt'>DNA hydroxymethylation in single cells</span><span style='font-size:10.5pt;font-family:"Arial","sans-serif"'> to gain new insights into </span><span style='font-size:10.5pt;font-family:"Arial","sans-serif"'>cues that guide differentiation during early mammalian development and into </span><span style='font-size:10.5pt;font-family:"Arial","sans-serif"'>mechanisms regulating heterogeneity within tumors.<o:p></o:p></span></p><p class=MsoNormal style='mso-margin-top-alt:auto;mso-margin-bottom-alt:auto'><o:p> </o:p></p><p class=MsoNormal style='text-align:justify'><span style='font-size:10.0pt;font-family:"Times New Roman","serif";color:black'><o:p> </o:p></span></p><p class=MsoNormal style='text-align:justify;background:white'><b><u><span style='font-size:14.0pt'><o:p><span style='text-decoration:none'> </span></o:p></span></u></b></p><p class=MsoNormal style='text-align:justify;background:white'><b><u><span style='font-size:14.0pt'><o:p><span style='text-decoration:none'> </span></o:p></span></u></b></p><p class=MsoNormal><span style='font-size:10.0pt'><o:p> </o:p></span></p><p class=MsoNormal style='text-align:justify;text-indent:27.35pt;line-height:15.0pt;mso-line-height-rule:exactly'><span style='font-family:"Arial","sans-serif"'><o:p> </o:p></span></p><p class=MsoNormal style='mso-margin-top-alt:auto;mso-margin-bottom-alt:auto'><span style='font-size:14.0pt'><o:p> </o:p></span></p><p class=MsoNormal style='mso-margin-top-alt:auto;mso-margin-bottom-alt:auto'><span style='font-size:14.0pt'><o:p> </o:p></span></p><p class=MsoNormal style='margin-right:2.25pt;text-align:justify;line-height:115%'><span style='font-size:14.0pt;line-height:115%'><o:p> </o:p></span></p><p class=MsoNormal><span style='color:#1F497D'><o:p> </o:p></span></p><p class=MsoNormal style='text-align:justify'><o:p> </o:p></p><p class=MsoNormal style='text-align:justify'><span style='color:#1F497D'><o:p> </o:p></span></p><p class=MsoNormal style='text-align:justify;text-indent:12.05pt'><span style='font-family:"Franklin Gothic Book","sans-serif"'><o:p> </o:p></span></p><p class=MsoNormal><o:p> </o:p></p><p class=MsoNormal><o:p> </o:p></p><p class=MsoNormal><o:p> </o:p></p></div></body></html>